Compositions and methods for affecting movement of contaminants, bodily fluids or other entities, and/or affecting other physiological conditions

ABSTRACT

Compositions which self-assemble under physiological conditions are formulated for application to wounds. The formulations include a pharmaceutically acceptable carrier or are provided as part of a medical device or coating. The formulations may also include other therapeutic, prophylactic or diagnostic agents. The formulation can be administered as appropriate for treatment of one or more disorders or conditions. For example, the formulation may be applied to repair an injury or during surgery of the lung, eye or dura, or following an epidural or spinal tap, to stop leakage of blood, interstitial fluid, or cerebrospinal fluid.

RELATED APPLICATIONS

This application claims priority under 35 U.S.C. §119(e) to U.S.Provisional Application No. 60/745,601, filed Apr. 25, 2006, thecontents of which are incorporated herein by reference.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT

This invention was made with government support under Grant No. EY00126,awarded by the National Institute of Health. The government has certainrights in this invention.

FIELD OF THE INVENTION

The present invention generally relates to compositions and methods foraffecting physiological conditions, including movement of bodily fluidsand/or contaminants.

BACKGROUND OF THE INVENTION

Despite the availability of blood products, blood loss is a major causeof morbidity and mortality. There are many causes of such loss,including severe injury and clinical conditions such as the rupture ofan aneurysm, esophageal or gastric ulcers, and esophageal varices. Aloss of integrity of a major artery can rapidly lead to death,particularly if it occurs in a setting where there is no rapid access tomedical care.

Bleeding during surgery is often a major concern. Blood loss can cause amyriad of problems for the patient while the presence of blood inundesirable locations can be detrimental to normal tissue or interferewith the surgeon's ability to view the operative field. The surgery mustbe delayed while blood is removed and the bleeding is brought undercontrol. Bleeding can be problematic even during minimally invasivesurgery (e.g., laparoscopic surgery). In some instances, surgeons mustconvert these preferred procedures into traditional open surgeries ifbleeding cannot be adequately controlled.

Bleeding can also be problematic in diagnostic and interventionalprocedures that involve the percutaneous introduction of instrumentationinto an artery, vein or smaller vessel. For example, procedures such ascoronaryangioplasty, angiography, atherectomy, and stenting of arteriesoften involve accessing the vasculature through a catheter placed into ablood vessel such as the femoral artery. Once the procedure is completedand the catheter or other instrument is removed, bleeding from thepunctured vessel must be controlled.

Options for controlling bleeding in any of these settings are limited.One of the oldest methods includes application of pressure, eitherdirectly to a vessel or to the body external to the vessel. Pressuremust be maintained until the bleeding is under control. This procedureis time-consuming and inconvenient, and the patient is at risk ofhematoma. Other physical methods include the use of clamps, clips,plugs, sponges, or the like. These devices have limited efficacy, andthey can be cumbersome to apply, particularly if there are many smallbleeding vessels. Use of heat to coagulate blood and cauterize bleedingvessels is widely used during surgery, but it is a destructive processthat can result in damage to collateral tissue. Furthermore, thesemethods require equipment and expertise and are thus not suitable foruse outside of medical settings. In addition to heat and mechanicaldevices, a variety of compounds have been used to promote hemostasis,but none of these are ideal.

Accordingly, improved methods are needed.

SUMMARY OF THE INVENTION

The present invention relates to structures positioned to inhibitmovement of a bodily fluid and/or contaminant of an animal comprising amaterial selected from among peptidomimetics, nucleotidomimetics,diblock copolymers, triblock copolymers, N-alkylacrylamides, oligomershaving a backbone capable of adopting a helical or sheet conformation,or a combination of more than one of these, wherein the material hasbeen applied to a portion of the animal prior to, during, and/or afterthe occurrence of a surgical incision or wound of the animal, in anamount sufficient to substantially inhibit movement of the bodily fluidand/or contaminant, and the structure comprises a product of interactionbetween the material and at least one ionic species present in or on theanimal, such that the structure substantially inhibits movement of thebodily fluid and/or contaminant of the animal.

The present invention also relates to compositions comprising a materialselected from among peptidomimetics, nucleotidomimetics, diblockcopolymers, triblock copolymers, N-alkylacrylamides, oligomers having abackbone capable of adopting a helical or sheet conformation, or acombination of more than one of these, wherein the material is able,upon interaction with at least one ionic species present in or on aanimal, to form a structure that substantially inhibits movement of abodily fluid and/or contaminant of the animal.

The present invention also relates to compositions comprising a materialselected from among peptidomimetics, nucleotidomimetics, diblockcopolymers, triblock copolymers, N-alkylacrylamides, oligomers having abackbone capable of adopting a helical or sheet conformation, or acombination of more than one of these, wherein the material has beenapplied to a surface of a component, comprising a medical device orother article, in an amount sufficient to substantially inhibit movementof a contaminant on the surface of the component relative to thecomponent, and wherein the material is able, upon interaction with atleast one ionic species present on the surface of the component, to forma structure that substantially inhibits movement of the contaminant.

The present invention also provides methods for inhibiting movement of abodily fluid and/or contaminant on or in an animal comprisingadministering a composition to a portion of an animal prior to, during,and/or after the occurrence of a surgical incision or wound of theanimal, in an amount sufficient to substantially inhibit movement of thebodily fluid and/or contaminant, wherein the composition comprises apeptidomimetic, nucleotidomimetic, diblock copolymer, triblockcopolymer, N-alkylacrylamide, oligomer having a backbone capable ofadopting a helical or sheet conformation, or a combination of more thanone of these; and allowing the composition to interact with at least oneionic species present in or on the animal to form a structure thatinhibits movement of the bodily fluid and/or contaminant.

The present invention also provides methods for inhibiting movement of acontaminant comprising applying a composition to a surface of acomponent, the component comprising a medical device or other article,in an amount sufficient to substantially inhibit movement of acontaminant on the surface of the component relative to the component,wherein the composition comprises a peptidomimetic, nucleotidomimetic,diblock copolymer, triblock copolymer, N-alkylacrylamide, oligomerhaving a backbone capable of adopting a helical or sheet conformation,or a combination of more than one of these; and allowing the compositionto interact with at least one ionic species present in or on the animalto form a structure that inhibits movement of the contaminant.

The present invention also provides methods for minimally invasivesurgery, comprising administering a composition to a portion of a animalprior to, during, and/or after the occurrence of a minimally-invasivesurgical procedure of the animal, in an amount sufficient tosubstantially inhibit movement of a bodily fluid and/or contaminant,wherein the composition comprises a peptidomimetic, nucleotidomimetic,diblock copolymer, triblock copolymer, N-alkylacrylamide, oligomerhaving a backbone capable of adopting a helical or sheet conformation,or a combination of more than one of these.

The present invention also provides methods for inhibiting movement of abodily fluid and/or contaminant on or in a animal, comprisingadministering, to a portion of an animal prior to, during, and/or afterthe occurrence of a surgical incision or wound of the animal, in anamount sufficient to substantially inhibit movement of the bodily fluidand/or contaminant, in combination: a composition comprising a material,selected from among peptidomimetics, nucleotidomimetics, diblockcopolymers, triblock copolymers, N-alkylacrylamides, oligomers having abackbone capable of adopting a helical or sheet conformation, or acombination of more than one of these, and a solution which, whencombined with the composition, dissolves or hydrates the materials toform a material capable of inhibiting movement of a bodily fluid and/orcontaminant.

Compositions including materials which self-assemble under physiologicalconditions may be formulated for application to wounds. Theconcentration of the self-assembling materials in any given formulationcan vary and can be between approximately 0.1% and 99%, inclusive,preferably between 0.1% and 10%. In one embodiment, the concentration ofthe self-assembling materials (e.g., in a liquid formulation) can beapproximately 0.1-3.0% (1-30 mg/ml) (e.g., 0.1-1.0%; 1.0-2.0%; 2.0-3.0%or 1.0-3.0%). The concentration of self-assembling materials can behigher in stock solutions and in solid (e.g., powdered) formulations.Solid preparations may have a concentration of self-assembling materialsapproaching 100% (e.g., the concentration of self-assembling materialscan be 95, 96, 97, 98, 99% or more (e.g., 99.99%) of the composition).Whether in liquid or solid form, the materials can be brought to thedesired concentration prior to use by addition of a pharmaceuticallyacceptable diluent (e.g. deionized water), fillers, or oil.

The formulations include a pharmaceutically acceptable carrier or areprovided as part of a medical device or coating. The formulations mayalso include other therapeutic, prophylactic or diagnostic agents. Theseinclude, but are not limited to, anti-inflammatories, vasoactive agents,anti-infectives, anesthetics, growth factors, and/or cells. Metals maybe added as chelators or to decrease adhesion. In one embodiment, theformulation is provided as a dry or lyophilized powder which can beadministered directly as a powder or a tablet, disc, or wafer whichhydrates at the site of application, or suspended or dissolved in aliquid, most preferably aqueous, and applied as a spray, paint, orinjection or a hydrogel including a material such as chitin, collagen,alginate, or synthetic polymer. In the preferred embodiment, thematerial is provided in combination with an oil, and forms a laminate.In another embodiment, the formulation is provided as a coating on adevice, for example a stent or a catheter, which may be applied bydissolving the self-assembling materials in an aqueous solution anddrying on the device, or mixed with a polymeric carrier and applied tothe device. In yet another embodiment, the formulation is provided in abandage, foam or matrix, in which the materials may be dispersed orabsorbed. The formulation could also be in the form of sutures, tape, oradhesive, or applied to a material such as a surgical drape, to preventcontamination. The material is also useful to isolate tissue, forexample, during removal of a specific tissue or tumor, in the eye orlung to prevent hemorrhage (as in response to hemorrhagic fever), forpreservation or stabilization of tissue for subsequent transplantationor reattachment, and as a bulking, stabilizing or hydrating agent. Incertain embodiments, the material may be useful in a blood substitute,since it does not lyse blood and inhibits platelet aggregation. Inanother embodiment, the materials, at concentrations insufficient forself assembly, can be used to stabilize blood.

One or more of the compositions described herein can be assembled inkits, together with instructions for use. For example, the kits caninclude a biocompatible composition including self-assembling materials(or a concentrated solution or powdered formulation thereof, togetherwith a diluent) and a vasoconstrictor, a coloring agent, or an analgesicor anesthetic agent and instructions for their combination (if notalready combined) and use (e.g., dilution and administration). The kitscan further include one or more of the additional agents describedherein. These agents can be present within the self assemblingcomposition or packaged separately, and they can include one or moretypes of biological cells, an antibiotic or other therapeutic, collagen,an anti-inflammatory agent, a growth factor, or a nutrient. The kit mayalso include one or more of a syringe (e.g., a barrel syringe or a bulbsyringe), a needle, a pipette, gauze, sponges, cotton, swabs, a bandage,a nosebleed plug, a disinfectant, surgical thread, scissors, a scalpel,a sterile fluid, a spray canister, including those in which a liquidsolution is sprayed through a simple hand pump, a sterile container, ordisposable gloves.

The formulation can be administered as appropriate for treatment of oneor more disorders or conditions, such as those noted above. For example,the formulation may be applied to repair an injury or during surgery ofthe lung, eye or dura, or following an epidural or spinal tap, to stopleakage of blood, interstitial fluid, or cerebrospinal fluid. Theformulations can also be applied to the intestines, the biliary tractand/or the urinary tract to stop the movement of fluid in or out of thetissue/organ. The formulation may be administered to a burn or ulcer,especially when formulated with anesthetics, anti-inflammatories, growthfactors, and anti-infectives, in the form of a foam, matrix or bandage,to stop bleeding or loss of interstitial fluid. The formulation may bedispersed in a suture or adhesive for administration at the time of oras released following suturing or gluing of a wound, thereby limitingbleeding, loss of tissue fluids, or other fluids such as those producedby parenchymal tissues such as the liver, pancreas, and gastrointestinaltract. The formulation may be applied to any site of bleeding, in abandage, gauze, sponge, or other material, for immediate control ofbleeding, or released later to control bleeding if the initial treatmentsuch as suturing or pressure is insufficient. Dried fabric, dehydratedfoams or hydrogels, or bandages containing the formulation may be partof first aid kits for treatment of injuries, for example, in war, ataccident sites, or clinics where rapid treatment may be required andstorage space is limited. In embodiments featuring bandages ordressings, the bandage or dressing can include a first layer ofsufficient shape and size to cover a wound or a substantial portionthereof (e.g., the most injured portion of the tissue or the areableeding most profusely). The first layer can have a top surface, abottom surface, and a perimeter that is, optionally, wholly or partiallycovered with an adhesive. A second layer of the bandage or dressing canbe detachably affixed to the bottom surface of the first layer,optionally excluding the perimeter or any part of the perimeter bearingadhesive, and can include a liquid or non-liquid composition (e.g., agel, paste, foam, cream, ointment, or powdered composition) includingself-assembling peptides. The composition will come in contact with thewound upon application of the bandage or dressing and is transferablefrom the bandage or dressing to the wound site upon removal of the firstlayer or the first and second layers. In simpler configurations, thecomposition including self-assembling molecules can be associated withthe bottom of the first layer (e.g., interior to the adhesiveperimeter), and the second layer can be omitted. In either case, eitherthe first and/or second layers can include a transparent window, throughwhich some or all of the underlying wound can be viewed. The compositionincluding the self-assembling materials can be added to the bandagebefore it is packaged or just before use. In another embodiment, theformulation may include a further physical barrier, such as a layer ofsilicone film, to prevent loss of fluid by drying, after the active flowof fluids has been stopped by application of the formulation. Theformulation may be applied as a hydrogel, laminate, or spray.

The liquid formulations may be provided in a syringe or pipette having abarrel containing a composition including self-assembling materials anda means for expelling the composition from an open tip of the syringe orpipette (e.g., a plunger or bulb). The syringe may consist of one ormore compartments, so that mixing of the self-assembling materials withone or more other agents occurs at the time of application. Thecompartments may also contain excipients such as a material forming ahydrogel or adhesive in one compartment and the self-assemblingmaterials in the other compartment. In another embodiment, onecompartment may contain lyophilized particles of self-assemblingmaterials, and another compartment may contain a solvent or solution todissolve or hydrate the materials, or mixed with other powders for dryapplication. The composition within the barrel can further include anyof the agents described herein (e.g., one or more of a vasoconstrictor,a coloring agent, an anesthetic or analgesic agent, an antibiotic orother therapeutic, collagen, an anti-inflammatory agent, a growthfactor, or a nutrient).

The liquid and powder compositions are stable, preferably for a periodgreater than one year, more preferably greater than two years and mostpreferably greater than three years.

DETAILED DESCRIPTION

The present invention provides compositions and techniques for affectingmovement of bodily fluids, e.g. blood, and/or movement of contaminantssuch as potentially infectious microorganisms. Fluids can be partiallyor fully contained to prevent loss, or contaminants can be isolated andinhibited from movement to sites ideally kept free of them, by materialsand techniques of the invention. Articles such as medical equipment canbe coated for inhibition of contaminant movement as well. Materials andtechniques of the invention can be used in a medical treatment setting,or in other settings. As will be seen from the variety of disclosureherein, other uses of materials of the invention also are provided.

In one set of embodiments, compositions of the invention can bepositioned to affect a physiological condition, such as movement of abodily fluid and/or contaminant. For example, material of the inventionmay be applied to a surface, in some cases, prior to, during, and/orafter the occurrence of a surgical incision or wound, in an amountsufficient to partially or substantially inhibit or prevent movement ofa bodily fluid and/or contaminant.

In some cases, compositions of the invention may compriseself-assembling materials. While materials that can self-assemble, andthereby be transformed in configuration (optionally in combination withother materials such as bodily fluid or tissue) into a condition wheremovement of fluid and/or contaminant is inhibited may be used in someembodiments, the materials do not necessarily need to self-assemble inorder to function in accordance with the invention. Materials typicallyinteract with themselves (intramolecular or intermolecular interactionsof self-assembly or the like), and/or interact with the environment inwhich they are placed (e.g. with a surface of a cell, another surface,with a bodily fluid, or the like), in order to function as describedherein. This principle is discussed more fully below. Some aspects ofthe invention are described in connection with “self-assembling” or“self-assembled” materials, and it is to be understood that whereverthis terminology is used, the materials referred to may, but need not,self-assemble in order to function per the invention. Reference is madeto “assembling” materials in some instances, and this is meant toencompass materials that self-assemble and/or undergo anothertransformation toward use in accordance with the invention. In somecases, a particular material can self-assemble in some environments, andin other environments (e.g., in the presence of an auxiliary componentsuch as blood cells not present in the first environment) may notself-assemble at all.

The materials may be useful in, for example, controlling leakage ofbodily fluids such as blood, interstitial fluid, and/or cerebrospinalfluid. In some cases, the invention provides compositions such as thesein association with a bandage, spray, coating, or powder. In someembodiments, compositions of the invention may be also be sufficientlyclear (e.g., optically clear) to allow a physician to see and workthrough the material.

Compositions of the invention can be provided in a form suitable foruse, and, in some cases, may not self-assemble or interact in a way soas to be transformed to affect movement of fluid or contaminant, orother use, as described herein, until exposed to bodily fluid, skin, oran ingredient containing a species that promotes a change in thecomposition relative to itself (self-assembly) or relative to othermaterial (e.g. cells). Many compositions of the material undergo changewhen exposed to ionic species, including ions present in levels inherentin bodily fluids or on skin. Exposure of these materials to ionic, orother, species to cause change (typically, an increase in viscosity oreven transformation from an essentially fluid to an essentially gel orsemi-solid form) can be caused by exposure to skin, to bodily fluid, orto another component prior to or during application of the material tothe site of use (e.g. a treatment site of an animal, or a medicaldevice), and/or inherently present on a device through a manufacturingprotocol.

Many compositions and classes of compositions suitable for use inconnection with the invention are described below. These and othermaterials can be readily selected by those of ordinary skill in the art.

I. Formulations

A. Materials

Peptidomimetics

One class of materials that can be used in connection with the inventionare peptidomimetics. Peptidomimetics, as used herein, refers tomolecules which mimic peptide structure. Peptidomimetics have generalfeatures analogous to their parent structures, polypeptides, such asamphiphilicity. Examples of such peptidomimetic materials are describedin Moore et al., Chem. Rev. 101(12), 3893-4012 (2001).

The term “peptide,” as used herein includes “polypeptide” and“oligopeptide” and refers to a string of at least two amino acidresidues linked together by covalent bonds (e.g., peptide bonds). Usefulpeptides can vary in length so long as they retain the ability toself-assemble to an extent useful for one or more of the purposesdescribed herein. Peptides having as few as two amino acid residues oras many as approximately 200 residues may be suitable, and thoserecognized to self-assemble typically have a length within this range(e.g., 8-200, 8-36, 8-24, 8-16, 12-20, 6-64, or 16-20 amino acidresidues). Depending on the context, “peptide” may refer to anindividual peptide or to a collection of peptides having the same ordifferent sequences. In addition, one or more of the amino acid residuesin a self-assembling peptide can be altered or derivatized by theaddition of a chemical entity such as an acyl group, a carbohydrategroup, a carbohydrate chain, a phosphate group, a farnesyl group, anisofarnesyl group, a fatty acid group, or a linker for conjugation orfunctionalization. Useful peptides can also be branched, in which casethey will contain at least two amino acid polymers, each of whichconsists of at least three amino acid residues joined by peptide bonds.The two amino acid polymers themselves are linked, but not by a peptidebond.

While the sequences of the peptides can vary, useful sequences includethose that convey an amphiphilic nature to the peptides (e.g., thepeptides can include approximately equal numbers of hydrophobic andhydrophilic amino acid residues), and the peptides can be complementaryand structurally compatible. Complementary peptides have an ability tointeract through ionic or hydrogen bonds that form between residues(e.g., hydrophilic residues) on adjacent peptides in a structure. Forexample, a given hydrophilic residue in a peptide can either hydrogenbond or ionically pair with a hydrophilic residue on an adjacentpeptide. Unpaired residues can be exposed to the solvent.Peptide-peptide interaction may also involve van der Waals forces orother forces that do not constitute covalent bonds. The peptides arestructurally compatible when they are capable of maintaining asufficiently constant intrapeptide distance to allow assembly andstructure formation. While the intrapeptide distance can vary, it can bequite small (e.g., less than about 4, 3, 2, or 1 Å). The intrapeptidedistance (e.g., an average of a representative number of distances) canbe larger than this, however. These distances can be calculated based onmolecular modeling or based on a simplified procedure that has beenpreviously reported (see U.S. Pat. No. 5,670,483).

Where peptides are used, without wishing to be bound by any particulartheory as to how they function in accordance with the invention, it issuggested that their side chains (or R groups) partition into two faces,a polar face with positively and/or negatively charged ionic sidechains, and a nonpolar face with side chains that are considered neutralor uncharged at physiological pH (e.g., the side chain of an alanineresidue or residues having other hydrophobic groups). The positivelycharged and negatively charged amino acid residues on the polar face ofone peptide can form complementary ionic pairs with oppositely chargedresidues of another peptide. These peptides may therefore be calledionic, self-complementary peptides. If the ionic residues alternate withone positively and one negatively charged residue on the polar face(−+−+−+−+), the peptides may be described as “modulus I;” if the ionicresidues alternate with two positively and two negatively chargedresidues (−−++−−++) on the polar face, the peptides are described as“modulus II;” if the ionic residues alternate with three positively andthree negatively charged residues (+++−−−+++−−−) on the polar face, thepeptides are describe as “modulus III;” if the ionic residues alternatewith four positively and four negatively charged residues(++++−−−++++−−−−) on the polar face, they are described as “modulus IV.”A peptide having four repeating units of the sequence EAKA may bedesignated EAKA 16-I, and peptides having other sequences may bedescribed by the same convention.

Peptide-based structures can be formed of heterogeneous mixtures ofpeptides (i.e., mixtures containing more than one type of peptideconforming to a given formula or to two or more of the formulas). Insome embodiments, each of the types of peptides in the mixture are ableto self-assemble alone. In other embodiments, one or more of each typeof peptide would not, alone, self-assemble but the combination ofheterogeneous peptides may self-assemble (i.e., peptides in the mixtureare complementary and structurally compatible with each other). Thus,either a homogeneous mixture of self-complementary and self-compatiblepeptides of the same sequence or containing the same repeating subunit,or a heterogeneous mixture of different peptides which are complementaryand structurally compatible to each other, can be used.

In summary, peptides useful in the manner described herein can have, orcan include, a sequence of alternating hydrophobic and hydrophilic aminoacid residues that are complementary and structurally compatible. Asnoted, the peptides can vary in length and can be a multiple of fourresidues. For example, the peptides can be at least eight amino acids inlength (e.g., eight or 10 amino acids), at least 12 amino acids inlength (e.g., 12 or 14 amino acids), or at least 16 amino acids inlength (e.g., 16, 18, 20, 22, or 24 amino acids). Peptides that are lessthan 100 amino acid residues long, more preferably less thanapproximately 50 amino acids in length, may assemble more readily.

Either or both ends of a given peptide can be modified. For example, thecarboxyl and/or amino groups of the carboxyl- and amino-terminalresidues, respectively can be protected or not protected. The charge ata terminus can also be modified. For example, a group or radical such asan acyl group (RCO—, where R is an organic group (e.g., an acetyl group(CH3CO—)) can be present at the N-terminus of a peptide to neutralize an“extra” positive charge that may otherwise be present (e.g., a chargenot resulting from the side chain of the N-terminal amino acid).Similarly, a group such as an amine group (NH2) can be used toneutralize an “extra” negative charge that may otherwise be present atthe C-terminus (e.g., a charge not resulting from the side chain of theC-terminal amino acid residue). Where an amine is used, the C-terminuswould bear an amide (—CONH2). The neutralization of charges on aterminus may facilitate self-assembly. One of ordinary skill in the artwill be able to select other suitable groups.

Structures for use in the invention can be formed that have varyingdegrees of stiffness or elasticity. The structures typically have a lowelastic modulus (e.g., a modulus in the range of 1-10 kPa as measured bystandard methods, such as in a standard cone-plate rheometer). Lowvalues may be preferable, as they permit structure deformation as aresult of movement, in response to pressure, in the event of cellcontraction. The desired stiffness of the composition can be dictated bythe tissue/organ to which the composition is to be applied. Thestiffness can be controlled in a variety of ways, including by changingthe length, sequence, and/or concentration of the precursor molecules(e.g., self-assembling peptides). Other methods for increasing stiffnesscan also be employed. For example, one can attach, to the precursors,biotin molecules or any other molecules that can be subsequentlycross-linked or otherwise bonded to one another. The molecules (e.g.,biotin) can be included at an N- or C-terminus of a peptide or attachedto one or more residues between the termini. Where biotin is used,cross-linking can be achieved by subsequent addition of avidin.Biotin-containing peptides or peptides containing other cross-linkablemolecules are within the scope of the present invention. For example,amino acid residues with aromatic rings may be incorporated andcross-linked by exposure to UV light. The extent of crosslinking can beprecisely controlled by applying the radiation for a predeterminedlength of time to peptides of known sequence and concentration. Theextent of crosslinking can be determined by light scattering, gelfiltration, or scanning electron microscopy using standard methods.Furthermore, crosslinking can be examined by HPLC or mass spectrometryanalysis of the structure after digestion with a protease, such asmatrix metalloproteases. Material strength may be determined before andafter cross-linking. Regardless of whether cross-linking is achieved bya chemical agent or light energy, the molecules may be cross-linked inthe course of creating a mold or when peptide-containing solutions areapplied to the body.

The half-life (e.g., the in vivo half-life) of the structures can alsobe modulated by incorporating protease or peptidase cleavage sites intothe precursors that subsequently form a given structure. Proteases orpeptidases that occur naturally in vivo or that are introduced (e.g., bya surgeon) can then promote degradation by cleaving their cognatesubstrates. Combinations of any of the modifications described here canbe made. For example, self-assembling peptides that include a proteasecleavage site and a cysteine residue and/or a cross-linking agent, kitsand devices containing them, and methods of using them can be utilized.

The peptide structures formed from any peptides, includingself-assembling peptides, made by any process can be characterized usingvarious biophysical and optical techniques, such as circular dichroism(CD), dynamic light scattering, Fourier transform infrared (FTIR),atomic force (tension) microscopy (ATM), scanning electron microscopy(SEM), and transmission electron microscopy (TEM). For example,biophysical methods can be used to determine the degree of beta-sheetsecondary structure in the peptide structure. Filament and pore size,fiber diameter, length, elasticity, and volume fraction can bedetermined using quantitative image analysis of scanning and/ortransmission electron micrographs. The structures can also be examinedusing several standard mechanical testing techniques to measure theextent of swelling, the effect of pH and ion concentration on structureformation, the level of hydration under various conditions, the tensilestrength, as well as the manner in which various characteristics changeover the period of time required for the structures to form and degrade.These methods allow one of ordinary skill in the art to determine whichof the various alternatives and peptides described herein are mostsuitable for use in the various methods, and allow optimization of thevarious processes.

Peptidomimetic materials of the invention can include (α-peptides,β-peptides, γ-peptides, and δ-peptides). Copolymers of these peptidescan also be used. Examples of α-peptide peptidomimetics include, but arenot limited to, N,N′-linked oligoureas, oligopyrrolinones,oxazolidin-2-ones, azatides and azapeptides.

Examples of suitable β-peptides include, but are not limited to,β-peptide foldamers, α-aminoxy acids, sulfur-containing β-peptideanalogues, and hydrazino peptides.

Examples of suitable γ-peptides include, but are not limited to,γ-peptide foldamers, oligoureas, oligocarbamates, and phosphodiesters.

Examples of suitable δ-peptides include, but are not limited to,alkene-based δ-amino acids and carbopeptoids, such as pyranose-basedcarbopeptoids and furanose-based carbopeptoids.

Oligomers Having Backbones which can Adopt Helical or SheetConformations

Another class of compounds useful in connection with the inventionincludes oligomers having backbones which can adopt helical or sheetconformations. Example of such compounds include, but are not limitedto, compounds having backbones utilizing bipyridine segments, compoundshaving backbones utilizing solvophobic interactions, compounds havingbackbones utilizing side chain interactions, compounds having backbonesutilizing hydrogen bonding interactions, and compounds having backbonesutilizing metal coordination.

Examples of suitable compounds containing backbones utilizing bipyridinesegments include, but are not limited to, oligo(pyridine-pyrimidines),oligo(pyridine-pyrimidines) with hydrazal linkers, andpyridine-pyridazines.

Examples of suitable compounds containing backbones utilizingsolvophobic interactions include, but are not limited to,oligoguanidines, aedamers (structures which take advantage of thestacking properties of aromatic electron donor-acceptor interactions ofcovalently linked subunits) such as oligomers containing1,4,5,8-naphthalene-tetracarboxylic diimide rings and1,5-dialkoxynaphthalene rings, and cyclophanes such as substitutedN-benzyl phenylpyridinium cyclophanes.

Examples of suitable compounds containing backbones utilizing side chaininteractions include, but are not limited to, oligothiophenes such asolihothiophenes with chiral p-phenyl-oxazoline side chains, andoligo(m-phenylene-ethynylene)s.

Examples of compound containing backbones utilizing hydrogen bondinginteractions include, but are not limited to, aromatic amide backbonessuch as oligo(acylated 2,2′-bipyridine-3,3′-diamine)s andoligo(2,5-bis[2-aminophenyl]pyrazine)s, diaminopyridine backbonestemplated by cyanurate, and phenylene-pyridine-pyrimidine ethynylenebackbones templated by isophthalic acid.

Examples of suitable compounds containing backbones utilizing metalcoordination include, but are not limited to, zinc bilinones,oligopyridines complexed with Co(II), Co(III), Cu(II), Ni(II), Pd(II),Cr(III), or Y(III), oligo(m-pheylene ethynylene)s containingmetal-coordinating cyano groups, and hexapyrrins.

Nucleotidomimetics

Another class of molecules which can be used in the invention, and canin some instances self assemble, are nucleotidomimetics such as isomericoligonucleotides, modified carbohydrates, nucleotides with modifiednucleotide linkages, and nucleotides with alternative nucleobases.

Examples of suitable isomeric nucleotides include, but are not limitedto, iso-RNA and iso-DNA and α-DNA (change in the anomeric configurationfrom β to α), alt-DNA, and 1-DNA.

Examples of suitable modified carbohydrates include, but are not limitedto, backbones with C 1′-bases connectivities such as tetrofuranosyloligonucleotides, pentopyranosyl oligonucleotides, and hexopyranosyloligonucleotides; backbones with C2′-base connectivities such asisonucleotides (repositioning of the base sugar connection from C 1 tothe C2 position), HNAs (insertion of an additional methylene groupbetween the 04′ and C 1′ position of a furanose), ANAs (incorporation ofa C3′-(S)-hydroxyl group), MNAs (inversion of the C3′-OH configurationfrom (S) in ANAs to (R)), CNAs (replacement of the O of the hexose witha methylene group), CeNAs (introduction of a 5′-6′ alkene within theanalogous ring), as well as other ring systems, torsionally restrictedoligonucleotides such as bicyclic oligonucleotides, LNAs (restriction ofthe pentofaranose backbone to the 3′-endo configuration), torsionallyflexible oligonucleotides such as base sugar extensions (insertion ofmethylene and ethylene groups into both α- and β-deoxynucleotides) andacyclic backbones (glycerol derivatives incorporating phosphodiesterlinkages).

Examples of nucleotides with modified nucleotide linkages include, butare not limited to, PNAs (peptide nucleic acids), NDPs(nucleo-δ-peptides), fused sugar-base backbones, and cationic linkages.

Examples of suitable alternative nucleobases include, but are notlimited to, nucleotides with alternative aromatic nucleobases.

Other Materials

Other materials which can self-assemble or can otherwise be useful inthe invention include N-alkylacrylamide oligomers and di- and triblockco-polymers. N-alkylacrylamides can assume self-assembled sheet-likestructures. Examples of suitable block copolymers includecopolypeptides, polypeptide-PEGS, PEO-polybutadienes,PEG-polysaccharides, etc.

B. Formation of Materials

Prior to use, materials of the invention may be maintained in neatcondition, or contained in (e.g., dissolved in) a solution that issubstantially free of ions (e.g., monovalent ions) or that contains asufficiently low concentration of ions to prevent significantself-assembly or other transformation (e.g., a concentration of ionsless than 10, 5, 1, or 0.1 mM). Self-assembly or other transformationtoward inhibition of movement of fluid or contaminant may be initiatedor enhanced at any subsequent time by the addition of an ionic solute ordiluent to a solution of the material or by a change in pH. For example,NaCl at a concentration of between approximately 5 mM and 5 M can inducethe assembly of macroscopic structures within a short period of time(e.g., within a few minutes). Lower concentrations of NaCl may alsoinduce assembly but at a slower rate. Alternatively, self-assembly orother transformation may be initiated or enhanced by introducing thematerials (whether dry, in a semi-solid gel, or dissolved in a liquidsolution that is substantially free of ions) into a fluid (e.g., aphysiological fluid such as blood or gastric juice) or an area (e.g., abody cavity such as the nose or mouth or a cavity exposed by a surgicalprocedure) comprising such ions. Generally, self-assembly occurs uponcontacting the material with such a solution in any manner.

A wide variety of ions, including anions and cations (whether divalent,monovalent, or trivalent), can be used to cause self-assembly or othertransformation of materials of the invention to a condition where theycan prevent movement of fluid and/or contaminant. For example, one canpromote a phase transition by exposure to monovalent cations such asLi+, Na+, K+, and Cs+. The concentration of such ions required to induceor enhance self-assembly or other transformation is typically at least 5mM (e.g., at least 10, 20, or 50 mM). Lower concentrations alsofacilitate assembly, although at a reduced rate. When desired,assembling materials can be delivered with a hydrophobic material (e.g.a pharmaceutically acceptable oil) in a concentration that permitsself-assembly or other transformation, but at a reduced rate. When suchmaterials are mixed with a hydrophobic agent such as an oil or lipid theassembly of the material forms different structures. The structures willappear like ice on a layer of oil. In some cases when another materialis added, the material will assemble into various other threedimensional structures that may be suitable for loading of a therapeuticagent. The hydrophilic part of the molecule will assemble in such a wayas to minimize hydrophobic-hydrophilic interaction, thereby creating abarrier between the two environments. Several experiments have shownthat the assembling materials will align on the surface of the oil likeice on water with the hydrophobic part of the molecule toward thesurface and the hydrophilic portion of the molecule facing away from theoil, or will form toroidal like structures with the hydrophobic materialcontained inside. This type of behavior enables the encapsulation oftherapeutics or other molecules of interest for delivery in the body.

Depending on the formulation and desired properties of the macroscopicstructure (e.g., the stiffness of the scaffold or the rate of itsformation), the concentration of precursors (e.g., self-assemblingmaterials) can vary from approximately 0.01% w/v (0.1 mg/ml) toapproximately 99.99% w/v (999.9 mg/ml), inclusive. For example, theconcentration prior to scaffold formation can be between approximately0.1% (1 mg/ml) and 10% (100 mg/ml), inclusive (e.g., about 0.1%-5%;0.5%-5%; 1.0%; 1.5%; 2.0%; 2.5%; 3.0%; or 4.0% or more). The precursors(e.g., self-assembling materials) can be formulated as powders andadministered in a powder form or resuspended. If dry, the materials canthen self-assemble following contact with bodily fluids (e.g., at a siteof injury).

The materials can be formed within regularly or irregularly-shapedmolds, which may include a body cavity or a portion of the body (e.g.,the lumen of a blood vessel) or which may be an inert material such asplastic or glass. The structures or scaffolds can be made to conform toa predetermined shape or to have a predetermined volume. To form astructure with a predetermined shape or volume (e.g., a desired geometryor dimension, including thin sheets or films), an aqueous solution ofthe material is placed in a pre-shaped casting mold, and the materialsare induced to self-assemble by the addition of a plurality of ions.

Alternately, ions may be added to the solution shortly before placingthe solution into the mold, provided that care is taken to place thesolution into the mold before substantial assembly occurs. Where themold is a tissue (e.g., the lumen of a blood vessel or othercompartment, whether in situ or not), the addition of an ionic solutionmay not be necessary. The resulting characteristics of the material, thetime required for assembly, and the dimensions of the macroscopicstructure that is formed are governed by the concentration and amount ofsolution that is applied, the concentration of ions used to induceassembly of the structure, and the dimensions of the casting apparatus.The scaffold can achieve a gel-like or substantially solid form at roomtemperature, and heat may be applied to facilitate the molding (e.g.,one can heat a solution used in the molding process (e.g., aprecursor-containing solution) to a temperature ranging up to about bodytemperature (approximately 37° C.). Once the scaffold has reached thedesired degree of firmness, it can be removed from the mold and used fora purpose described herein.

Materials that assemble and/or undergo a phase transition (e.g., atransition from a liquid state to a semi-solid, gel, etc.) when theycome in contact with the body are useful in preventing the movement ofbodily substances. Self-assembly or phase transition is triggered bycomponents found in a subject's body (e.g., ions) or by physiological pHand is assisted by physiological temperatures. Self-assembly or phasetransition can begin when the compositions are exposed to or broughtinto contact with a subject's body and may be facilitated by the localapplication of heat to the area where the composition has been (or willbe) deposited. Based on studies to date, self-assembly occurs rapidlyupon contact with internal bodily tissues without the application ofadditional heat. The time required for effective assembly and/or phasetransition can occur in 60 seconds or less following contact with asubject's internal tissues or to conditions similar to those foundwithin the body (e.g., in 50, 40, 30, 20, or 10 seconds or less). Insome circumstances, such as where the concentration of self-assemblingagents in the composition is low or where the movement of the bodilysubstance is substantial, self-assembly or phase transition may takelonger to achieve the desired effect, for example, up to a minute, 5minutes, 10 minutes, 30 minutes, an hour, or longer. For example, asolution containing a self-assembling peptide applied to sites of bloodvessel transection in the brain, liver, or muscle provided completehemostasis within times as short as 10 seconds following application.Ion-containing solutions may be preferred when the compositions are usedto protect a subject from contamination, as phase transitions do notoccur, or do not readily occur, when non-ionic compositions contactintact skin.

The compositions can form structures that are substantially rigid (e.g.,solid or nearly solid) or that assume a definite shape and volume (e.g.,structures that conform to the shape and volume of the location to whicha liquid composition was administered, whether in vivo or ex vivo). Thesolidified material may be somewhat deformable or compressible afterassembly or phase transition, but will not substantially flow from onearea to another, as compositions at a different point along the liquidto solid continuum may do, which may be due, at least in part, to theirability to undergo phase transitions. As a result, the compositions canbe used to prevent the movement of a bodily substance in a subject inneed thereof. Self-assembly can also be achieved ex vivo by exposure toconditions within a certain range of physiological values (e.g.,conditions appropriate for cell or tissue culture). While liquidformulations are readily dispensed, the compositions administered mayalso be in a gel form that may become stiffer upon contact with thesubject's body.

The concentration of the self-assembling materials in any givenformulation can vary and can be between approximately 0.1% (1 mg/ml) and10% (100 mg/ml), inclusive. For example, the concentration of theself-assembling peptides (e.g., in a liquid formulation) can beapproximately 0.13.0% (1-30 mg/ml) (e.g., 0.1-1.0%; 1.0-2.0%; 2.0-3.0%or 1.0-3.0%). The concentration of self-assembling materials can behigher in stock solutions and in solid (e.g., powdered) formulations. Insolid preparations, the concentration of self-assembling materials canapproach 100% (e.g., the concentration of self-assembling peptides canbe 95, 96, 97, 98, 99% or more (e.g., 99.99%) of the composition).Whether in liquid or solid form, the materials can be brought to thedesired concentration prior to use by addition of a diluent (e.g.,deionized water), powder, wetting agent, or a therapeutic, diagnostic orprophylactic agent.

Regardless of the precise nature of the self-assembling materials, uponexposure to conditions such as those described herein, the materials canform membranous two- or three-dimensional structures including a stablemacroscopic porous matrix having ordered interwoven nanofibers (e.g.,fibers approximately 10-20 nm in diameter, with a pore size of about50-100 nm in a linear dimension). Three-dimensional macroscopic matricescan have dimensions large enough to be visible under low magnification(e.g., about 10 fold or less), and the membranous structures can bevisible to the naked eye, even if transparent. Althoughthree-dimensional, the structures can be exceedingly thin, including alimited number of layers of molecules (e.g., 2, 3, or more layers ofmolecules). Typically, each dimension of a given structure will be atleast 10 μm in size (e.g., two dimensions of at least 100-1000 μm insize (e.g., 1-10 mm, 10-100 mm, or more)). The relevant dimensions maybe expressed as length, width, depth, breadth, height, radius, diameter,or circumference in the case of structures that have a substantiallyregular shape (e.g., where the structure is a sphere, cylinder, cube, orthe like) or an approximation of any of the foregoing where thestructures do not have a regular shape.

The self-assembling materials can form a hydrated material whencontacted with water under conditions such as those described herein(e.g., in the presence of a sufficient concentration (e.g.,physiological concentrations) of ions (e.g., monovalent cations)). Thematerials may have a high water content (e.g., approximately 95% or more(e.g., approximately 97%, 98%, 99% or more)), and the compositions canbe hydrated but not substantially self-assembled. A given value may be“approximate” in recognition of the fact that measurements can varydepending, for example, on the circumstances under which they are madeand the skill of the person taking the measurement. Generally, a firstvalue is approximately equal to a second when the first falls within 10%of the second (whether greater than or less than) unless it is otherwiseclear from the context that a value is not approximate or where, forexample, such value would exceed 100% of a possible value.

The properties and mechanical strength of the structures or scaffoldscan be controlled as required through manipulation of the componentstherein. For example, the stiffness of an assembled gel can be increasedby increasing the concentration of self-assembling materials therein.The sequences, characteristics, and properties of the materials and thestructures formed by them upon self-assembly are discussed furtherbelow.

The compositions can be formulated as concentrated stocks or in dryform, and these can be diluted or dissolved to form biocompatiblecompositions, which are substantially non-toxic to biological cells invitro or in vivo. For example, the compositions can contain materials inquantities that do not elicit a significant deleterious effect on therecipient's body (e.g., a prohibitively severe immunological orinflammatory reaction, or unacceptable scar tissue formation).

When a solution containing non-assembled materials is laid down on abiological tissue, the materials having sufficient proximity to thetissue assemble, causing the solution to gel. Any solution that remainsdistant from the tissue remains liquid, as the self-assembling materialshave not yet been exposed to conditions that promote their assembly. Asthe material is disturbed (e.g., by performing a surgical procedure),liquid material appears to gel as it comes into sufficient contact withthe body. At times, the compositions can take on characteristics rangingfrom a liquid to those of a solid, appearing gel- or salve-like or as aslurry).

The formulations disclosed herein are stable for a period of timegreater than one year, preferably greater than two years, and morepreferably greater than three years.

Many aspects of the invention are described in the context of“self-assembly”, or “self-assembling” materials. As noted above, it isto, be understood that, in some aspects and settings, compositions ofthe invention can self-assemble into a material that may have adifferent viscosity, phase, morphology, or other difference in form thatallows it to serve one of the uses of the invention such as inhibitionof movement of bodily fluids. However, self-assembly is not required inall aspects. For example, in some settings, compositions of theinvention may serve to interact with cells, tissues, or other componentsin a way that the benefits of the invention are realized withoutself-assembly but through another procedure, such as immobilizing cellsrelative to each other or to tissue, immobilizing different tissuecomponents relative to each other, or other interactions with otherinventive compositions or components of a subject to be treated, device,or article, in a manner such that goals of the invention are realized.Thus, while self-assembly may play an important role in certain aspectsof the invention and certain settings, it is not a necessary aspect ofall embodiments of the invention.

C. Additional Therapeutic, Prophylactic and Diagnostic Agents

The formulations typically include an excipient or otherpharmaceutically acceptable carrier or are provided as part of a medicaldevice or coating. The formulations may also include other therapeutic,prophylactic or diagnostic agents. In a preferred embodiment, these maybe anti-inflammatories, vasoactive agents, anti-infectives, anesthetics,growth factors, and/or cells.

These can be peptides or proteins, polysaccharides or saccharides,nucleic acids nucleotides, proteoglycan, lipid, carbohydrate, or a smallmolecule, typically an organic compound, having multiple carbon-carbonbonds that may be isolated from nature or prepared via chemicalsynthesis. Small molecules have relatively low molecular weights (e.g.,less than about 1500 g/mol) and are not peptides or nucleic acids. Thesubstance can also be a biomolecule, which is a molecule such as apeptide, protcoglycan, lipid, carbohydrate, or nucleic acid havingcharacteristics typical of molecules found in living organisms. Likesmall molecules, biomolecules can be naturally occurring or may beartificial (i.e., they may be molecules that have not been found innature). For example, a protein having a sequence that has not beenfound in nature (e.g., one that does not occur in a publicly availabledatabase of sequences) or that has a known sequence modified in anunnatural way by a human hand (e.g., a sequence modified by altering apost-translational process such as glycosylation) is an artificialbiomolecule. Nucleic acid molecules encoding such proteins (e.g., anoligonucleotide, optionally contained within an expression vector) arealso biomolecules and can be incorporated into the compositionsdescribed herein. For example, a composition can include a plurality ofself-assembling materials and cells that express, or that are engineeredto express, a protein biomolecule (by virtue of containing a nucleicacid sequence that encodes the protein biomolecule).

Many different therapeutic, prophylactic or diagnostic agents can beincorporated into the formulation. Representative vasoconstrictorsinclude epinephrine and phenylephrine; representative coloring agentsinclude arsenazo III, chlorophosphonazo III, antipyrylazo 111, murexide,Eriochrome Black T, Eriochrome Blue SE, oxyacetazo I, carboxyazo III,tropolone, methylthymol blue, and Mordant Black 32; representativeanesthetic agents include benzocaine, bupivacaine, butamben picrate,chloroprocaine, cocaine, curate, dibucaine, dyclonine, etidocaine,lidocaine, mepivacaine, pramoxine, prilocaine, procaine, propoxycaine,ropivacaine, tetracaine, or combinations thereof. Local application ofthe anesthetic agent may be all that is required in some situations, forexample, for a burn or other wound to the skin, including decubitusulcers, or for minimally invasive surgeries. Combining local anestheticswith the self-assembling materials, whether combined by virtue of beingpresent in the same formulation or by virtue of co-administration, canhelp contain the anesthetic within the body and reduce the amountentering the circulation.

Vasoconstrictors such as phenylephrine can be included to prolong theeffect of local anesthesia (e.g., 0.1-0.5% phenylephrine). Analgesicagents other than a local anesthetic agent, such as steroids,non-steroidal anti-inflammatory agents like indomethacin, plateletactivating factor (PAF) inhibitors such as lexipafant, CV 3988, and/orPAF receptor inhibitors such as SR163-441. An anti-infective orantimicrobial agent (e.g., an antibiotic, antibacterial, antiviral, orantifungal agent) can be included for either systemic or localadministration. Examples include (β-lactam antibiotics such aspenicillins and cephalosporins and other inhibitors of cell wallsynthesis such as vancomycin, chloramphenicol, tetracyclines,macrolides, clindamyin, streptogramins, aminoglycosides, spectinomycin,sulfonamides, trimethoprim, quinolones, amphotericin B, flucytosine,azoles such as ketoconazole, itraconazole, fluconazole, clotrimazole,and miconazole, griseofulvin, terbinafine, and nystatin. Theantimicrobial can be topically administered (e.g., to treat skininfections or burns, or to help prevent infection at a site of catheterinsertion (e.g., an intravenous catheter), for example, kanamycin,neomycin, bacitracin, polymixin, topical sulfonamides such as mafenideacetate or silver sulfadiazine, or gentamicin sulfate. The antimicrobialcan also be a broad spectrum agent. For example, a second, third, orfourth generation cephalosporin can be used. These agents may be activeagainst a wide range of bacteria including both gram positive and gramnegative species. Such antibacterial agents may be particularlyappropriate where the present scaffolds are used to inhibit movement ofintestinal contents such as during intestinal resection or other surgerythat purposefully or accidentally disturbs the integrity of theintestinal wall. One of ordinary skill in the art will be able to selectappropriate antimicrobial agents by considering factors such as thepatient's history (e.g., any history of an allergic reaction to suchagents), the location to which the peptides are to be applied, the typeof infectious agent likely to be present, and so forth. Any of thecompositions described herein, whether they contain only self-assemblingprecursors or precursors and one or more bioactive molecules (andwhether in a liquid, semi-solid, or solid farm), can include a coloringagent. Suitable coloring agents include commercially available foodcolorings, natural and synthetic dyes, and fluorescent molecules.

Preferably, the coloring agent is nontoxic or is included at such lowconcentrations as to minimize any toxic effect. The use of a coloringagent allows for improved visualization of an area that is covered by astructure or scaffold and can facilitate removal, if such removal isdesired. The coloring agent can be one that changes color when it comesinto contact with a contaminated area (e.g., a color change may betriggered by the contamination itself (e.g., by the blood or bacteriapresent at a wound site)). For example, a metabolic product of abacterium may trigger a color change. Conditions such as pH or redoxstate induced by contaminants may also be detected. Exemplary indicatorsinclude arsenzazo III, chlorophosphonazo III, antipyrylazo III,murexide, Eriochrome Black T and Eriochrome Blue SE for Mg2+, oxyacetazoI, carboxyazo III, tropolone, methylthymol blue, and Mordant Black 32.AlamarBlue, a redox indicator, and phenol red are also of use in thecompositions and methods.

Many other active agents can be included in the compositions. Forexample, a number of growth factors can be included to accelerate one ormore aspects of healing (e.g., angiogenesis, cell migration, processextension, and cell proliferation). These types of compositions can be“included” as others can, by virtue of inclusion in the compositions orby virtue of co-administration in the present methods. Examples includevascular endothelial growth factor (VEGF), a transforming growth factor(TGF) such as transforming growth factor p, a platelet derived growthfactor (PDGF), an epidermal growth factor (EGF), a nerve growth factor(NGF), an insulin-like growth factor (e.g., insulin-like growth factorI), a glial growth factor (GGF), a fibroblast growth factor (FGF), etc.It will be appreciated that in many cases these terms refer to a varietyof different molecular species. For example, several transforming growthfactor R species are known in the art. One of ordinary skill in the artwill be guided in the selection of an appropriate growth factor byconsidering, for example, the site at which the composition is to beadministered. For example, an EGF can be included in compositionsapplied to the skin; an NGF and/or GGF can be included in compositionsapplied to nerves or the nervous system; and so forth.

The growth factor or another agent can be a chemotactic substance, whichhas the ability, in vivo or in cell culture, to recruit cells to a siteat which the substance is present. The cells recruited may have thepotential to contribute to the formation of new tissue or to repairexisting, damaged tissue (e.g., by contributing structurally and/orfunctionally to the tissue (e.g., by providing growth factors orcontributing to a desirable immune response)). Certain chemotacticsubstances can also function as proliferation agents (e.g., neurotropicfactors such as NGF or BDNF).

The compositions can also be used in combination with or instead ofcompounds such as cyanoacrylates, oxidized cellulose, fibrin sealants,collagen gel, thrombin powder, microporous polysaccharide powders,clotting factors (e.g., Factor V, Factor VIII, fibrinogen, orprothrombin) and zeolite powders.

In one embodiment, vitamins may be added to the material such as vitaminK after liver surgery. In addition, other vitamins can be added tofacilitate the reconstruction of tissue or skin when applied topicallyin combination with the material. This could be after injury or in thenormal course of topical hydration.

The one or more therapeutic, diagnostic and/or prophylactic agents canbe administered simultaneously with the self-assembling materials in thesame formulation, administered simultaneously in separate formulations,or sequentially.

It will be understood that therapeutic molecules are generallyadministered in an effective amount in order to achieve a clinicallysignificant result, and effective dosages and concentrations are knownin the art. These dosages and concentrations can guide the selection ofdosages and concentrations in the present context. Bioactive moleculescan be provided at a variety of suitable concentrations and in suitableamounts (e.g., in the microgram or milligram range, or greater). Forguidance, one can consult texts such as Goodman and Gilman's ThePharmacological Basis of Therapeutics, 10th Ed., and Katzung, Basic andClinical Pharmacology.

Cells

Where cells are delivered to a patient (e.g., to promote tissuehealing), utologous cells can be used. In one embodiment, the cellscould be hematopoietic cells from the patient, dispersed in the materialand implanted. In another embodiment, the cells can be cord red bloodcells.

Molded scaffolds as described above, liquid compositions, gels, solids(e.g. powders) or other semi-solid embodiments may include one or moreadditional substances such as bioactive molecules or cells. In someinstances, the cell may secrete the bioactive molecule either naturallyor following genetic engineering (e.g., to express and/or secrete arecombinant protein). The structures described herein are able tosupport cell attachment, viability, and growth; these have been observedwhen cells are cultured on the surface of the material or when cellsgrow within the material (e.g., when encapsulated). In addition, thestructures are able to serve as substrates for neurite growth andsynapse formation when neurons are grown on or within them. Thus,bioactive molecules and cells can be encapsulated within the peptidestructures and maintain substantial function and viability when soencapsulated (see, e.g., U.S. Ser. Nos. 09/778,200 and 10/196,942).

D. Excipients, Carriers, and Devices

The formulations include a pharmaceutically acceptable carrier or areprovided as part of a medical device or coating.

In one embodiment, the formulation is provided as a dry or lyophilizedpowder which can be administered directly as a powder which hydrates atthe site of application, or suspended or dissolved in a liquid, mostpreferably aqueous, and applied as a spray, paint, or injection or ahydrogel such as chitin, collagen, alginate, or synthetic polymer. Inanother embodiment, the formulation is administered as a compressedwafer, disc, or tablet. In still another embodiment, the formulation isprovided as a coating on a device, for example a stent or a catheter,which may be dissolved in an aqueous solution and dried on the device,or mixed with a polymeric carrier and applied to the device. In yetanother embodiment, the formulation is provided in a bandage, foam ormatrix, in which the peptides may be dispersed or absorbed. Theformulation could also be in the form of sutures, tape, or adhesive.

Conventionally, local anesthetics are delivered by topicaladministration (e.g., formulated as an ointment, cream, or solution) orinjected into an area where the nerve fibers one wishes to block reside.The formulation may be administered to a burn or ulcer, especially whenformulated with anesthetics, antiinflammatories, growth factors, andantiinfectives, in the form of a foam, matrix or bandage, to stopbleeding or loss of interstitial fluid.

One or more of the compositions described herein can be assembled inkits, together with instructions for use. For example, the kits caninclude a biocompatible composition including self-assembling peptides(or a concentrated solution or powdered formulation thereof, togetherwith a diluent) and a vasoconstrictor, a coloring agent, or an analgesicor anesthetic agent and instructions for their combination (if notalready combined) and use (e.g., dilution and administration). The kitscan further include one or more of the additional agents describedherein. These agents can be present within a peptide-based compositionor packaged separately, and they can include one or more types ofbiological cells, an antibiotic or other therapeutic, collagen, ananti-inflammatory agent, a growth factor, or a nutrient. The kit mayalso include one or more of a syringe (e.g., a barrel syringe or a bulbsyringe), a needle, a pipette, gauze, sponges, cotton or the like,swabs, a bandage, a nosebleed plug, a disinfectant, surgical thread,scissors, a scalpel, a sterile fluid, a spray canister, including thosein which a liquid solution is sprayed through a simple hand pump, asterile container, or disposable gloves.

The formulation can be administered as appropriate for treatment of oneor more disorders. For example, the formulation may be applied to repairan injury or dealing surgery of the lung or dura, or following anepidural or spinal tap, to stop leakage of cerebrospinal fluid. Theformulation may be dispersed in a suture or adhesive for administrationat the time of or as released following suturing or gluing of a wound,thereby limiting bleeding, loss of tissue fluids, or other fluids suchas those produced by parenchymal tissues such as the liver, pancreas,and gastrointestinal tract. The formulation may be applied to any siteof bleeding, in a bandage, gauze, sponge, or other material, forimmediate control of bleeding, or released later to control bleeding ifthe initial treatment such as suturing or pressure is insufficient.

Dried fabric, dehydrated foams or hydrogels, or bandages containing theformulation may be part of first aid kids for treatment of injuries, forexample, in war, at accident sites, or clinics where rapid treatment maybe required and storage space is limited.

In some embodiments, compositions including self-assembling materialscan be associated with surgical sponges. For example, liquidcompositions can be drawn into commercially available sponges prior toor during their use. Studies indicate that hemostasis can besatisfactorily achieved without traditional sponges, but there may beinstances where including compositions containing a self-assemblingmaterial may be beneficial (e.g., where a patient is experiencingprofound bleeding or where the goal of treatment is temporarystabilization). The compositions employed can include any of thenon-fibrous agents described herein. The sponges can be any known in theart, including woven and non-woven sponges and those designedspecifically for dental or ophthalmic surgeries. See, e.g., U.S. Pat.Nos. 4,098,728; 4,211,227; 4,636,208; 5,180,375; and 6,711,879.

In embodiments featuring bandages or dressings, the bandage or dressingcan include a first layer of sufficient shape and size to cover a woundor a substantial portion thereof (e.g., the most injured portion of thetissue or the area bleeding most profusely). The first layer can have atop surface, a bottom surface, and a perimeter that is, optionally,wholly or partially covered with an adhesive. A second layer of thebandage or dressing can be detachably affixed to the bottom surface ofthe first layer, optionally excluding the perimeter or any part of theperimeter bearing adhesive, and can include a liquid or non-liquidcomposition (e.g., a gel, paste, foam, cream, ointment, or powderedcomposition) including self-assembling peptides. The composition willcome in contact with the wound upon application of the bandage ordressing and is transferable from the bandage or dressing to the woundsite upon removal of the first layer or the first and second layers. Insimpler configurations, the composition comprising self-assemblingmaterials can be associated with the bottom of the first layer (e.g.,interior to the adhesive perimeter), and the second layer can beomitted. In either case, either the first and/or second layers caninclude a transparent window, through which some or all of theunderlying wound can be viewed. The composition including theself-assembling materials can be added to the bandage before it ispackaged or just before use. In another embodiment, the formulation mayinclude a further physical barrier, such as a layer of silicon film, toprevent loss of fluid by drying, after the active flow of fluids hasbeen stopped by application of the formulation.

The formulations may also be administered as immediate or controlledrelease formulations. A delayed release dosage form is one that releasesa drug (or drugs) at a time other than promptly after administration. Anextended release dosage form is one that allows at least a twofoldreduction in dosing frequency as compared to the drug presented as aconventional dosage form (e.g. as a solution or prompt drug-releasing,conventional solid dosage form). A modified release dosage form is onefor which the drug release characteristics of time, course and/orlocation are chosen to accomplish therapeutic or convenience objectivesnot offered by conventional dosage forms such as solutions, ointments,or promptly dissolving dosage forms. Delayed release and extendedrelease dosage forms and their combinations are types of modifiedrelease dosage forms.

Matrix forming materials are materials which form strong, viscous gelsupon hydration and provide control of drug diffusion and release. Inhydrophilic matrix systems, matrix forming materials are uniformlyincorporated throughout the tablet. Upon contact with water, the outertablet layer is partially hydrated, forming a gel layer. The rate ofdiffusion of the drug(s) out of the gel layer and the rate of erosion ofthe gel layer determine overall tablet dissolution and drug deliveryrates. Examples of matrix forming materials include cellulose ethersthat are water-soluble such as methylcellulose, ethyl cellulose andhydroxypropyl methylcellulose.

Formulations are prepared using a pharmaceutically acceptable “barrier”composed of materials that are considered safe and effective and may beadministered to an individual without causing undesirable biologicalside effects or unwanted interactions. The “carrier” is all componentspresent in the pharmaceutical formulation other than the activeingredient or ingredients. The term “carrier” includes but is notlimited to diluents, binders, lubricants, disintegrators, fillers,matrix-forming compositions and coating compositions.

“Carrier” also includes all components of the coating composition whichmay include plasticizers, pigments, colorants, stabilizing agents, andglidants. The delayed release dosage formulations may be prepared asdescribed in references such as “Pharmaceutical dosage form tablets”,eds. Liberman et al. (New York, Marcel Dekker, Inc., 1989),“Remington—The science and practice of pharmacy”, 20th ed., LippincottWilliams & Wilkins, Baltimore, Md., 2000, and “Pharmaceutical dosageforms and drug delivery systems”, 6th Edition, Ansel et al., (Media,Pa.: Williams and Wilkins, 1995) which provides information on carriers,materials, equipment and processes for preparing tablets and capsulesand delayed release dosage forms of tablets, capsules, and granules.

Examples of suitable coating materials include, but are not limited to,cellulose polymers such as cellulose acetate phthalate, hydroxypropylcellulose, hydroxypropyl methylcellulose, hydroxypropyl methylcellulosephthalate and hydroxypropyl methylcellulose acetate succinate; polyvinylacetate phthalate, acrylic acid polymers and copolymers, and methacrylicresins that are commercially available under the trade name Eudragit™(Roth Pharma, Westerstadt, Germany), Zein, shellac, and polysaccharides.Additionally, the coating material may contain conventional carrierssuch as plasticizers, pigments, colorants, glidants, stabilizationagents, pore formers and surfactants. Optional pharmaceuticallyacceptable excipients present in the drug-containing tablets, beads,granules or particles include, but are not limited to, diluents,binders, lubricants, disintegrants, colorants, stabilizers, andsurfactants.

Diluents, also termed “fillers,” are typically necessary to increase thebulk of a solid dosage form so that a practical size is provided forcompression of tablets or formation of beads and granules. Suitablediluents include, but are not limited to, dicalcium phosphate dihydrate,calcium sulfate, lactose, sucrose, mannitol, sorbitol, cellulose,microcrystalline cellulose, kaolin, sodium chloride, dry starch,hydrolyzed starches, pre-gelatinized starch, silicone dioxide, titaniumoxide, magnesium aluminum silicate and powder sugar.

Binders are used to impart cohesive qualities to a solid dosageformulation, and thus ensure that a tablet or bead or granule remainsintact after the formation of the dosage forms. Suitable bindermaterials include, but are not limited to, starch, pre-gelatinizedstarch, gelatin, sugars (including sucrose, glucose, dextrose, lactoseand sorbitol), polyethylene glycol, waxes, natural and synthetic gumssuch as acacia, tragacanth, sodium alginate, cellulose, includinghydroxypropylmethylcellulose, hydroxypropylcellulose, ethylcellulose,and veegum, and synthetic polymers such as acrylic acid and methacrylicacid copolymers, methacrylic acid copolymers, methyl methacrylatecopolymers, aminoalkyl methacrylate copolymers, polyacrylicacid/polymethacrylic acid and polyvinylpyrrolidone. Some of thematerials which are suitable as binders can also be used asmatrix-forming materials such as hydroxypropyl methyl cellulose, ethylcellulose, and microcrystalline cellulose. Lubricants are used tofacilitate tablet manufacture. Examples of suitable lubricants include,but are not limited to, magnesium stearate, calcium stearate, stearicacid, glycerol behenate, polyethylene glycol, talc, and mineral oil.

Disintegrants are used to facilitate dosage form disintegration or“breakup” after administration, and generally include, but are notlimited to, starch, sodium starch glycolate, sodium carboxymethylstarch, sodium carboxymethylcellulose, hydroxypropyl cellulose,pre-gelatinized starch, clays, cellulose, alginine, gums or cross linkedpolymers, such as cross-linked PVP (Polyplasdone™ XL from GAF ChemicalCorp).

Stabilizers are used to inhibit or retard drug decomposition reactionswhich include, by way of example, oxidative reactions.

Surfactants may be anionic, cationic, amphoteric or nonionic surfaceactive agents. Suitable anionic surfactants include, but are not limitedto, those containing carboxylate, sulfonate and sulfate ions. Examplesof anionic surfactants include sodium, potassium, ammonium salts of longchain alkyl sulfonates and alkyl aryl sulfonates such as sodiumdodecylbenzene sulfonate; dialkyl sodium sulfosuccinates, such as sodiumdodecylbenzene sulfonate; dialkyl sodium sulfosuccinates, such as sodiumbis-(2-ethylthioxyl)-sulfosuccinate; and alkyl sulfates such as sodiumlauryl sulfate. Cationic surfactants include, but are not limited to,quaternary ammonium compounds such as benzalkonium chloride,benzethonium chloride, cetrimonium bromide, stearyl dimethylbenzylammonium chloride, polyoxyethylene and coconut amine. Examples ofnonionic surfactants include ethylene glycol monostearate, propyleneglycol myristate, glyceryl monostearate, glyceryl stearate,polyglyceryl-4-oleate, sorbitan acylate, sucrose acylate, PEG-150laurate, PEG400 monolaurate, polyoxyethylene monolaurate, polysorbates,polyoxyethylene octylphenylether, PEG-1000 cetyl ether, polyoxyethylenetridecyl ether, polypropylene glycol butyl ether, Poloxamer™ 401,stearoyl monoisopropanolamide, and polyoxyethylene hydrogenated tallowamide. Examples of amphoteric surfactants include sodiumN-dodecyl-β-alanine, sodium N-lauryl-β-iminodipropionate,myristoamphoacetate, lauryl betaine and lauryl sulfobetaine.

If desired, the tablets, beads, granules or particles may also containminor amount of nontoxic auxiliary substances such as wetting oremulsifying agents, dyes, pH buffering agents, and preservatives.

Extended release formulations are generally prepared as diffusion orosmotic systems, for example, as described in “Remington—The science andpractice of pharmacy” (20th ed., Lippincott Williams & Wilkins,Baltimore, Md., 2000). A diffusion system typically consists of twotypes of devices, a reservoir and a matrix, and is well known anddescribed in the art. The matrix devices are generally prepared bycompressing the drug with a slowly dissolving polymer carrier into atablet form. The three major types of materials used in the preparationof matrix devices are insoluble plastics, hydrophilic polymers, andfatty compounds. Plastic matrices include methyl acrylate-methylmethacrylate, polyvinyl chloride, and polyethylene.

Hydrophilic polymers include cellulosic polymers such as methyl andethyl cellulose, hydroxyalkylcelluloses such as hydroxypropyl-cellulose,hydroxypropylmethylcellulose, sodium carboxymethylcellulose, andCarbopol™ 934, polyethylene oxides and mixtures thereof. Fatty compoundsinclude, but are not limited to, various waxes such as carnauba wax andglyceryl tristearate and wax-type substances including hydrogenatedcastor oil or hydrogenated vegetable oil, or mixtures thereof. Incertain embodiments, the plastic material is a pharmaceuticallyacceptable acrylic polymer, including but not limited to, acrylic acidand methacrylic acid copolymers, methyl methacrylate, methylmethacrylate copolymers, ethoxyethyl methacrylates, cyanoethylmethacrylate, aminoalkyl methacrylate copolymer, poly(acrylic acid),poly(methacrylic acid), methacrylic acid alkylamine copolymerpoly(methyl methacrylate), poly(methacrylic acid) (anhydride),polymethacrylate, polyacrylamide, poly(methacrylic acid anhydride), andglycidyl methacrylate copolymers. In certain embodiments, the acrylicpolymer is comprised of one or more ammonio methacrylate copolymers.Ammonio methacrylate copolymers are well known in the art, and aredescribed in NF XVII as fully polymerized copolymers of acrylic andmethacrylic acid esters with a low content of quaternary ammoniumgroups.

Alternatively, extended release formulations can be prepared usingosmotic systems or by applying a semi-permeable coating to the dosageform. In the latter case, the desired drug release profile can beachieved by combining low permeable and high permeable coating materialsin suitable proportion.

An immediate release portion can be added to the extended release systemby means of either applying an immediate release layer on top of theextended release core using a coating or compression process or in amultiple unit system such as a capsule containing extended and immediaterelease beads. Extended release tablets containing hydrophilic polymersare prepared by techniques commonly known in the art such as directcompression, wet granulation, or dry granulation. Their formulationsusually incorporate polymers, diluents, binders, and lubricants as wellas the active pharmaceutical ingredient. The usual diluents includeinert powdered substances such as starches, powdered cellulose,especially crystalline and microcrystalline cellulose, sugars such asfructose, mannitol and sucrose, grain flours and similar edible powders.Typical diluents include, for example, various types of starch, lactose,mannitol, kaolin, calcium phosphate or sulfate, inorganic salts such assodium chloride and powdered sugar. Powdered cellulose derivatives arealso useful.

Typical tablet binders include substances such as starch, gelatin andsugars such as lactose, fructose, and glucose. Natural and syntheticgums, including acacia, alginates, methylcellulose, andpolyvinylpyrrolidone can also be used. Polyethylene glycol, hydrophilicpolymers, ethylcellulose and waxes can also serve as binders. Alubricant is necessary in a tablet formulation to prevent the tablet andpunches from sticking in the die. The lubricant is chosen from suchslippery solids as talc, magnesium and calcium stearate, stearic acidand hydrogenated vegetable oils. Extended release tablets containing waxmaterials are generally prepared using methods known in the art such asa direct blend method, a congealing method, and an aqueous dispersionmethod. In the congealing method, the drug is mixed with a wax materialand either spray-congealed or congealed and screened and processed.

The preferred coating weights for particular coating materials may bereadily determined by those skilled in the art by evaluating individualrelease profiles for tablets, beads and granules prepared with differentquantities of various coating materials. It is the combination ofmaterials, method and form of application that produce the desiredrelease characteristics, which one can determine only from the clinicalstudies. The coating composition may include conventional additives,such as plasticizers, pigments, colorants, stabilizing agents, glidants,etc. A plasticizer is normally present to reduce the fragility of thecoating, and will generally represent about 10 wt. % to 50 wt. %relative to the dry weight of the polymer. Examples of typicalplasticizers include polyethylene glycol, propylene glycol, triacetin,dimethyl phthalate, diethyl phthalate, dibutyl phthalate, dibutylsebacate, triethyl citrate, tributyl citrate, triethyl acetyl citrate,castor oil and acetylated monoglycerides. A stabilizing agent ispreferably used to stabilize particles in the dispersion. Typicalstabilizing agents are nonionic emulsifiers such as sorbitan esters,polysorbates and polyvinylpyrrolidone. Glidants are recommended toreduce sticking effects during film formation and drying, and willgenerally represent approximately 25 wt. % to 100 wt. % of the polymerweight in the coating solution. One effective glidant is talc. Otherglidants such as magnesium stearate and glycerolmonostearates may alsobe used. Pigments such as titanium dioxide may also be used. Smallquantities of an anti-foaming agent, such as a silicone (e.g.,simethicone), may also be added to the coating composition.

Both non-biodegradable and biodegradable matrices can be used fordelivery of the self-assembling peptides, although biodegradablematrices are preferred. These may be natural or synthetic polymers,although synthetic polymers are preferred due to the bettercharacterization of degradation and release profiles. The polymer isselected based on the period over which release is desired. In somecases linear release may be most useful, although in others a pulserelease or “bulk release” may provide more effective results. Thepolymer may be in the form of a hydrogel (typically in absorbing up toabout 90% by weight of water), and can optionally be crosslinked withmultivalent ions or polymers.

Representative synthetic polymers that can be used for delivery includepolyamides, polycarbonates, polyalkylenes, polyalkylene glycols,polyalkylene oxides, polyalkylene terepthalates, polyvinyl alcohols,polyvinyl ethers, polyvinyl esters, polyvinyl halides,polyvinylpyrrolidone, polyglycolides, polysiloxanes, polyurethanes andco-polymers thereof, alkyl cellulose, hydroxyalkyl celluloses, celluloseethers, cellulose esters, nitro celluloses, polymers of acrylic andmethacrylic esters, methyl cellulose, ethyl cellulose, hydroxypropylcellulose, hydroxy-propyl methyl cellulose, hydroxybutyl methylcellulose, cellulose acetate, cellulose propionate, cellulose acetatebutyrate, cellulose acetate phthalate, carboxylethyl cellulose,cellulose triacetate, cellulose sulphate sodium salt, poly(methylmethacrylate), poly(ethyl methacrylate), poly(butylmethacrylate),poly(isobutyl methacrylate), poly(hexylmethacrylate), poly(isodecylmethacrylate), poly(lauryl methacrylate), poly(phenyl methacrylate),poly(methyl acrylate), poly(isopropyl acrylate), poly(isobutylacrylate), poly(octadecyl acrylate), polyethylene, polypropylene,poly(ethylene glycol), poly(ethylene oxide), poly(ethyleneterephthalate), poly(vinyl alcohols), poly(vinyl acetate, poly vinylchloride, polystyrene and polyvinylpyrrolidone.

Examples of non-biodegradable polymers include ethylene vinyl acetate,poly(meth)acrylic acid, polyamides, copolymers and mixtures thereof.Examples of biodegradable polymers include synthetic polymers such aspolymers of lactic acid and glycolic acid, polyanhydrides,poly(ortho)esters, polyurethanes, poly(butic acid), poly(valeric acid),and poly(lactide-co-caprolactone), and natural polymers such as alginateand other polysaccharides including dextran and cellulose, collagen,chemical derivatives thereof (substitutions, additions of chemicalgroups, for example, alkyl, alkylene, hydroxylations, oxidations, andother modifications routinely made by those skilled in the art), albuminand other hydrophilic proteins, zein and other prolamines andhydrophobic proteins, copolymers and mixtures thereof. In general, thesematerials degrade either by enzymatic hydrolysis or exposure to water invivo, by surface or bulk erosion.

Bioadhesive polymers of particular interest include bioerodiblehydrogels described by H. S. Sawhney, C. P. Pathak and J. A. Hubell inMacromolecules, 1993, 26, 581-587, polyhyaluronic acids, casein,gelatin, glutin, polyanhydrides, polyacrylic acid, alginate, chitosan,poly(methyl methacrylates), poly(ethyl methacrylates),poly(butylmethacrylate), poly(isobutyl methacrylate),poly(hexylmethacrylate), poly(isodecyl methacrylate), poly(laurylmethacrylate), poly(phenyl methacrylate), poly(methyl acrylate),poly(isopropyl acrylate), poly(isobutyl acrylate), and poly(octadecylacrylate).

The matrix can be in the form of microparticles such as microspheres,where peptides are dispersed within a solid polymeric matrix ormicrocapsules, where the core is of a different material than thepolymeric shell, and the peptide is dispersed or suspended in the core,which may be liquid or solid in nature. Unless specifically definedherein, microparticles, microspheres, and microcapsules are usedinterchangeably. Alternatively, the polymer may be cast as a thin slabor film, ranging from nanometers to four centimeters, a powder producedby grinding or other standard techniques, or even a gel such as ahydrogel. The polymer can also be in the form of a coating or part of astent or catheter, vascular graft, or other prosthetic device.

The matrices can be formed by solvent evaporation, spray drying, solventextraction and other methods known to those skilled in the art.

Bioerodible microspheres can be prepared using any of the methodsdeveloped for making microspheres for drug delivery, for example, asdescribed by Mathiowitz and Langer, J. Controlled Release 5, 13-22(1987); Mathiowitz, et al., Reactive Polymers 6, 275-283 (1987); andMathiowitz, et al., J. Appl. Polymer Sci. 35, 755-774 (1988). Theselection of the method depends on the polymer selection, the size,external morphology, and crystallinity that is desired, as described,for example, by Mathiowitz, et al., Scanning Microscopy 4, 329-340(1990); Mathiowitz, et al., J. Appl. Polymer Sci. 45, 125-134 (1992);and Benita, et al., J. Pharm. Sci. 73, 1721-1724 (1984). In solventevaporation, described for example, in Mathiowitz, et al., (1990),Benita, and U.S. Pat. No. 4,272,398 to Jaffe, the polymer is dissolvedin a volatile organic solvent. The peptide either in soluble form ordispersed as fine particles, is added to the polymer solution, and themixture is suspended in an aqueous phase that contains a surface activeagent such as poly(vinyl alcohol). The resulting emulsion is stirreduntil most of the organic solvent evaporates, leaving solidmicrospheres. In general, the polymer can be dissolved in methylenechloride. Microspheres with different sizes (1-1000 microns) andmorphologies can be obtained by this method which is useful forrelatively stable polymers such as polyesters and polystyrene. However,labile polymers such as polyanhydrides may degrade due to exposure towater. For these polymers, hot melt encapsulation and solvent removalmay be preferred.

In hot melt encapsulation, the polymer is first melted and then mixedwith the solid particles of peptides. The mixture is suspended in anon-miscible solvent such as silicon oil and, with continuous stirring,heated to 5° C. above the melting point of the polymer. Once theemulsion is stabilized, it is cooled until the polymer particlessolidify. The resulting microspheres are washed by decantation withpetroleum ether to give a free-flowing powder. Microspheres withdiameters between one and 1000 microns can be obtained with this method.The external surface of spheres prepared with this technique are usuallysmooth and dense. This procedure is useful with water labile polymers,but is limited to use with polymers with molecular weights between 1000and 50000. Solvent removal was primarily designed for use withpolyanhydrides. In this method, the drug is dispersed or dissolved in asolution of a selected polymer in a volatile organic solvent likemethylene chloride. The mixture is then suspended in oil, such assilicon oil, by stirring, to form an emulsion. Within 24 hours, thesolvent diffuses into the oil phase and the emulsion droplets hardeninto solid polymer microspheres. Unlike solvent evaporation, this methodcan be used to make microspheres from polymers with high melting pointsand a wide range of molecular weights.

Microspheres having a diameter between one and 300 microns can beobtained with this procedure. The external morphology of the spheres ishighly dependent on the type of polymer used. In spray drying, thepolymer is dissolved in methylene chloride (0.04 g/ml). A known amountof active drug is suspended (if insoluble) or co-dissolved (if soluble)in the polymer solution. The solution or the dispersion is thenspray-dried. Double walled microspheres can be prepared according toU.S. Pat. No. 4,861,627 to Mathiowitz.

Hydrogel microspheres made of gel-type polymers such as alginate orpolyphosphazines or other dicarboxylic polymers can be prepared bydissolving the polymer in an aqueous solution, suspending the materialto be incorporated into the mixture, and extruding the polymer mixturethrough a microdroplet forming device, equipped with a nitrogen gas jet.The resulting microspheres fall into a slowly stirring, ionic hardeningbath, as described, for example, by Salib, et al., PharmazeutischeIndustrie 40-11A, 1230 (1978). Chitosan microspheres can be prepared bydissolving the polymer in acidic solution and crosslinking withtripolyphosphate. For example, carboxymethylcellulose (CMC) microsphereare prepared by dissolving the polymer in an acid solution andprecipitating the microspheres with lead ions. Alginate/polyethyleneimide (PEI) can be prepared to reduce the amount of carboxyl groups onthe alginate microcapsules.

Other delivery systems including films, coatings, pellets, slabs, anddevices can be fabricated using solvent or melt casting, and extrusion,as well as standard methods for making composites. The polymer can beproduced by first mixing monomers and peptides as described by Sawhney,et al., and polymerizing the monomers with UV light. The polymerizationcan be carried out in vitro as well as in vivo.

E. Devices for Administration

The liquid formulations may be provided in a syringe or pipette having abarrel containing a composition including self-assembling peptides and ameans for expelling the composition from an open tip of the syringe orpipette (e.g., a plunger or bulb). The syringe may consist of one ormore compartments, so that mixing of the self-assembling materials withone or more other agents occurs at the time of application. Thecompartments may also contain an excipient such as a material forming ahydrogel or adhesive in one compartment and the self-assemblingmaterials in the other compartment. In another embodiment, onecompartment may contain lyophilized powder or particles ofself-assembling peptides, and another compartment may contain solutionto dissolve or hydrate the peptides, or other powders to mix with theself assembling materials for dry application. The composition withinthe barrel can further include any of the agents described herein (e.g.,one or more of a vasoconstrictor, a coloring agent, an anesthetic oranalgesic agent, an antibiotic or other therapeutic, collagen, ananti-inflammatory agent, a growth factor, or a nutrient).

The self-assembling material can be applied as a coating by spraying ordipping the device into the material, the material can be impregnatedinto a bandage, gauze or other absorbent material, the material can bemixed with a polymeric material.

II. Methods of Administration

Materials of the invention may be administered in any suitable setting.In some embodiments, the material may be used in a medical treatmentsetting. “Medical treatment setting,” as used herein, would beunderstood to those of ordinary skill in the art and means a clinicalsetting such as a hospital, clinic, doctor's office, surgical theater,or a mobile clinical setting such as a battlefield trauma treatmentcenter, or the like. Medical treatment setting also includes treatmentoutside of the technically clinical setting, such as home treatment,involving medical supplies such as bandages, gauze, and other materialand related techniques for the treatment of wounds and/or prevention ofadverse effects associated with wounds or contamination. Medicaltreatment setting is to be distinguished from other setting such as theuse of household cleaners and related products, diapers, and like. Ofcourse, it is to be understood that certain aspects of the inventioninvolve use or treatment in a medical treatment setting, but in otheraspects the invention can be used outside of this setting in essentiallyany other suitable setting.

Any of the agents described herein, including cells, therapeutic,prophylactic or diagnostic compounds such as antibiotics and growthfactors can be introduced into a solution of the self assemblingmaterial prior to self-assembly in vitro or in vivo, and pre-moldedstructures that include one or more of these agents, optionally packagedin sterile material and/or provided with instructions for use. Thematerial can be used prophylactically or as a treatment in the absenceof additional agents. The bioactive agents can be approximately evenlydistributed throughout the scaffold or concentrated in one area oranother (e.g., on or near the surface, within a core area, gradedthroughout the scaffold or a region thereof, or layered therein (e.g.,concentrated in layers or evenly or unevenly distributed)). To achievean approximately even distribution of the substance within thestructure, one can mix the precursor-containing solution and thesubstance, which may also be in solution, prior to initiation ofself-assembly.

A. Sites of Administration

The material can be applied to a variety of different surfaces toprevent or control fluid passage or to function as a barrier. The amountof self-assembling agent is determined in part by the function of thematerial in controlling fluid flow, as well as the properties of anyother materials or structures associated with the self-assemblingmaterial, alone or in combination with other bioactive materials. Theself-assembling materials can be used to stop the movement of fluids inor out of tissues/organs.

The materials can be used to coat stents or enable the insertion intothe location with a minimum of complication and without causing unduemovement of bodily fluid or substances solid, liquid, or semisolid suchas a lipid. These materials can be incorporated into a stent orcatheters to facilitate placement while minimizing disruption or to thelocal tissue.

The materials can be incorporated into bandages as a dry powder, liquid,or slurry either before placement or during the manufacturing process.In some embodiments the material will be deposited in multiple layersthat will the assembly of a bandage in place that could have awaterproof outer layer with a self assembled material that will stop theflow of bodily fluid from a wound or may incorporate additional agentsto facilitate healing of the site.

In a first embodiment, the material is used to prevent or controlbleeding. The material may be applied as a powder, liquid, a gel, or aspart of a substrate such as a bandage or membrane. This may be appliedto a blood vessel, either within the lumen, for example at the time ofangioplasty, administered by or as a coating on a stent or catheter, orexterior to the vessel, typically at the site of anastomosis. Thematerial may be applied to tissues before, during or after surgery, toprevent bleeding, which is especially problematic with tissue such asliver, kidney or spleen, or other surgeries where there is a high riskof transfusion, or to seal and protect a tissue, for example, which isfor transplantation or reattachment.

The material is also particularly well suited to use in the eye toprevent hemorrhage or bleeders within the vitreous humor. Othersurgeries where the material should be beneficial include cornealtransplants, conjunctival surgery and glaucoma surgery. Other surgerieswhere the material should be beneficial include corneal transplants,conjunctival surgery, glaucoma surgery and refractive eye surgery (e.g.,Lasik in which the material can be applied to or around the cornealflap). The material is particularly advantageous during surgery since itis clear and the surgery is able to see through the material, as heoperates.

The material can be used to stop the flow of fluids other than blood.The material can be applied to burns to stop leakage of interstitialfluid. The material can be applied to the dura or lung as a dural orlung sealant. In one embodiment the material can be used to repair alung after a puncture wound, thereby restoring its ability to function.

Materials of the invention can also be added to burn wounds for severalpurposes. Burn wounds that are not open can be treated to immobilizecontaminants relative to the wound, i.e., to inhibit or preventinfection of the wound if and when it opens. Open burn wounds can betreated to similarly inhibit or prevent contamination, and/or to inhibitmovement of fluid relative to the wound, e.g., inhibit movement ofbodily fluid out of the wound, or external fluid into the wound.

The material can also be utilized in general oral surgery,periodontistry, and general dentistry, both as a barrier and to controlor prevent bleeding.

The use of the material in individuals with impaired coagulation(hemophilia, von Willebrands, vitamin K, protein S or protein Cdeficiency, 10 fulminant hepatitis, disseminated intravascularcoagulation (“DIC”), hemolytic-uremic syndrome (“HUS”)) is also animportant utility since the mechanism of action is independent of thenormal coagulation pathway. In another embodiment, the material isapplied, typically by spraying or injection, to the exterior of a tissuesuch as a tumor, to prevent breakage or metastasis at the time ofsurgery. The material controls bleeding during tumor resection, as wellas limits metastatis. This also minimizes the immune response that canbe caused by a laser during tumor resection. The material is also usefulin holding loose tumors together so that nothing is left behind whenthey are resected. There are several types of tumors that arenotoriously hard to resect because they are not held together tightly(i.e. they are not solid masses). The material is expected to beparticularly useful in tumor resection in the brain, and may be usefulin a dose-response manner for subcutaneous tumor resection. This maymake it easier to resect melanomas in the skin because it appears thatthe material also facilitates skin healing. The material can alsoinclude a marker reactive with certain types of antigens on the tumorcell surface, producing a calorimetric change to show that all of thecells have been removed or there are more that need to be resected. Theaddition of an indicator to the material as well as the ability of thematerial to act like a bio-barrier could reduce the need for second andthird operations as well as complications due to outside contaminationinto the surgical field. The material can also be used to delivermaterials such as DNA to the site of injury for an extended period oftime in vitro and for multiple treatments in vivo. Another advantage ofthe material is that it can be injected and gel in place, so that thematerial can be applied and reapplied during surgery, as necessary.

In still another embodiment, the material is particularly well suited tofunctioning as a barrier to prevent contamination, either to the tissueor from the tissue, for example, during intestinal surgery. The materialmay be applied to prepare an internal site prior to surgery, especiallysites such as the sinus cavities, and for surgeries such astransurethral and transvaginal surgery. The material should also beparticularly useful in cardiovascular surgery, where both barrier andhemostasis properties can be of value, for example, for heart valvepatients who are prone to adverse consequences such as valve ringabcesses (coat valve, add antibiotic), endocarditis (coat valve), aorticroot dissection (provide immediate hemostasis).

The material in combination with a metal such as silver hasanti-adhesive properties and can inhibit angiogenesis. Accordingly, itmay be useful in decreasing scarring and adhesions. The material isapplied after surgery, or to an injury such as a burn, to decreasescarring, fluid loss, and limit infection. This has further applicationin plastic surgery, especially for protection of areas cleaned anddebrided prior to closure or skin transplant, for example, inabdominoplasty, face lifts, flap donor sites, latissimus dorsi forbreast reconstruction.

In still another embodiment, the material is administered as a slurrythat can be drunk by a patient to reduce stomach bleeding, for example,from an ulcer, or decrease acidity. Alternatively, the material could beprovided as an enema to treat hemorrhoids or to fill in diverticula.

In yet another embodiment, the material can be used for a fertilitytreatment, preservation of eggs, and repair of scarred fallopian tubes.

The material may also be useful as a blood substitute, and as an organpreservation or stabilization material. For example, the materials maybe applied to and encapsulate a bodily fluid or component (e.g.,tissues, organs, skin, blood, and the like) to reduce damage prior to,during, and after transfer of the bodily fluid or component from thesubject. The material may preserve viability of the bodily fluid orcomponent for storage, transport, transplantation, and/orreimplantation.

As assembly is not irreversible, contained substances can be released.For example, the molecules or cells can be released from the structuresin vivo (e.g., small molecules can diffuse away and larger molecules andcells can be released as the structures degrade).

In still another embodiment, the material is used as a neuroprotectiveto minimize damage and scarring following neural injury. Peptide-basedstructures promote repair and regeneration of neural tissue (e.g., whenself-assembling peptides are applied to a lesion in the brain asdescribed in U.S. Ser. No. 10/968,790). The small size of the fiberswithin the scaffolds and/or the open “weave” structure of the materialspermits extension of cell processes and allows adequate diffusion ofnutrients and waste products in a manner that provides unique advantagesfor neural tissue regeneration.

In the course of promoting wound repair, the compositions may not onlyimprove the final outcome (e.g., reduced scar formation resulting in anoutcome that more closely resembles the original tissue), but alsoreduce the time required for healing. These results could not have beenpredicted on the basis of the results achieved following application tothe injured central nervous system, given the substantial differencesbetween neural and non-neural tissues.

Finally, the materials could be used as “nanodrapes” to prevent crosscontamination. For example, the materials could be applied as a coatingto the outside of the body and then induced to self assemble. Theself-assembled material may stop the movement of liquids into the body,the reducing the possibility of cross contamination.

B. Effective Dosages

In general, the amount of material required will vary depending onvarious factors such as the size or extent of an injury (which can, inturn, be expressed in terms of the length of an incision, the caliber ornumber of damaged blood vessels, the degree of a burn, the size anddepth of an ulcer, abrasion, or other injury). The amount may vary, forexample, from a few microliters to several milliliters or more, e.g.,tens or hundreds of milliliters. The device used to deliver the materialwill vary in accordance with the amount. For example, a syringe can beconveniently used to deliver smaller amounts, whereas a tube orsqueezable bottle would be more suitable for larger amounts. Aneffective amount (whether in reference to a scaffold, precursorsthereof, or another bioactive molecule present in the formulation),means the amount necessary to elicit an improved or desired biologicalresponse.

As will be appreciated by those of ordinary skill in this art, theeffective amount of an agent may vary depending on such factors as thedesired biological endpoint, the agent to be delivered, the nature ofthe site to which the agent is delivered, and the nature of thecondition for which the agent is administered. For example, an effectiveamount of a composition for accelerating hemostasis may be an amountsufficient to decrease the amount of blood lost between the time thatbleeding begins and the time when bleeding ends by at least 25% relativeto the amount of blood lost following treatment with cold saline or notreatment. An effective amount of a composition for acceleratinghemostasis may also be an amount sufficient to decrease the timerequired to achieve cessation of visible bleeding by at least 25%relative to the time required following treatment with cold saline or notreatment. An effective amount of a composition for promoting woundhealing may be an amount sufficient to decrease the time required toachieve a predetermined percent reduction in the size of a lesion by atleast 25% relative to the time required in the absence of suchtreatment. The amount of the composition provided can vary depending onthe severity of the subject's condition and should be sufficient toinhibit the unwanted movement to an extent that benefits the subject.The bodily substance can be blood, cerebrospinal fluid, pus, serousexudate, bile, pancreatic juice, or a substance normally containedwithin the gastrointestinal tract (e.g., the stomach or intestine), orurinary tract.

Those of ordinary skill in the art will understand the meaning of“amount sufficient” of “effective amount” in the context of use ofmaterials and techniques of the invention. Those of ordinary skill inthe art will also, from the description of materials and classes ofmaterials described herein, be able to easily select materials for use.At the outset, simple screening tests can be used to determine suitablematerials, and sufficient amounts of material, in a variety ofprocedures. For example, screening tests involving animals inanticipation of use in humans. Also, a screening test involving a small,test incisions, and treatment of the test incision prior to formation ofa larger incision, or the like (this screening test is generallydesirable for determining sufficient amount, after suitable material hasbeen identified through other means). Another screening test to identifysuitable materials involves establishing a test fluid having ioniccharacter similar to that of a bodily fluid or skin, and exposingmaterial to the fluid, or to an article with characteristics similar tothe skin. Through initial use, either in test settings or initial use inthe intended setting, amounts of material sufficient for a particularintended purpose can be readily determined by those of ordinary skill inthe art. It is also noted that some or all materials used in theinvention are non-toxic and can be used in excess with no adverseeffects. Accordingly, in many settings such as surgical procedures totreat incisions toward inhibiting bleeding, treatment of surfaces ofskin to prevent infection prior to incision or in the context of a burnwound, or the like, the use of a significant excess of material, andremoval of that excessive material if desired and where it is determinedthat sufficient material has been applied (via observation of theeffective use of the material) can be simply removed easily and withoutharm.

C. How Administered

The composition can be provided on the surface of the subject's bodyand/or provided within a cavity generated by force (e.g., by unexpectedtrauma or a surgical procedure). In this way the unwanted movement ofbodily substances can be inhibited in the context of a wide range ofsituations, including traumatic injury, a medical condition (e.g., achronic or prolonged medical condition associated with bleeding), orsurgical procedures (e.g., orthopedic surgery, dental surgery, cardiacsurgery, ophthalmic surgery, or plastic or reconstructive surgery). Forexample, where the unwanted movement of the bodily substance is theresult of trauma, the subject may have a partly or completely severedbody part, a laceration, abrasion, puncture wound, or a burn. Where thecompositions are applied to a surface of the body, they may not onlyinhibit the unwanted movement of a bodily substance, but also helpprotect the subject from contamination. For example, applying aself-assembling agent to the skin will impede the movement of anunwanted foreign substance on the skin or hair into a wound. When theunwanted movement of the bodily substance results from a chronic medicalcondition, the subject may experience recurrent bleeding. For example,the subject may be experiencing bleeding in connection with varicoseveins, including telangiectases, hemorrhoids, bleeding in the lungs(due, for example, to lung cancer, bronchitis, or a bacterial or viraldisease, including pneumonia or influenza), or esophageal varices.Medical conditions associated with recurrent bleeding can be treatedwith the compositions described herein, including those that containself-assembling peptides and a vasoconstrictor (e.g., phenylephrine,which can constitute about 0.25-0.5% of the composition). Where bleedingoccurs in the orophamyx or lungs, the compositions can be administeredthrough a metered dose inhaler. If the patient's condition hasdeteriorated to the point where artificial ventilation is required, thecompositions may be administered through a respirator or by lavage.

The unwanted movement of the bodily substance can also take place duringa surgical procedure, and that procedure can involve an incision withinthe subject's nervous system, eye, ear, nose, mouth, pharynx,respiratory system, cardiovascular system, digestive system, urinarysystem, reproductive system, musculoskeletal system, liver, orintegument. The methods can be carried out regardless of whether or notthe movement of the bodily substance was intentional. The compositionsdescribed herein can be applied before or after the unwanted movementoccurs (e.g., during a surgical procedure before the intentionaltransection of a blood vessel or after an unintentional transection of ablood vessel). For example, the surgical procedure can be carried outwith the intent to repair an aneurysm, impede bleeding within the brain,to treat esophageal varices, to treat an ulcer or to inhibit the loss ofgastric contents or intestinal contents (e.g., from a swollen orruptured appendix). The surgical procedure can involve resecting aportion of the subject's intestine. Other procedures that can be carriedout with the assistance of compositions including self-assembling agentsinclude arteriography, cardiac catheterization, insertion of a stent,assistance with a natural birth or birth by Caesarean section,hysterectomy, organ transplant, joint replacement, or excision of anintervertebral disk. These procedures are representative. The surgicalprocedure can be performed with the assistance of an endoscope orlaparoscope, and the compositions can be delivered independently or froma chamber situated within these devices and connected to a distal end bya passage for release onto the subject's tissues. Where the patient hasan ulcer, that ulcer can be an esophageal, gastric, duodenal, diabetic,or decubitus ulcer. More generally, the compositions can be applied toany disrupted area of the skin, and any of the methods described hereincan include a step of identifying a patient in need of treatment.

A self-assembling peptide nanofiber scaffold (SAPNS) can provide atransparent environment for the surgical field, while also creating anoptically clear liquid that allows operation through the resultantliquid and gel mix. The surgical field is often obscured with blood anddebris during an operation. In addition, clearing debris from thesurgical field usually requires irrigating the site with saline. Salineis only a temporary solution and needs to be continuously applied tomaintain a clear surgical field. This poses several issues: anycontamination in existence will easily spread; a small opening willrequire alternating between irrigation and operating; and duringintestinal operations use of saline can result in a massive infectionleading to post-operative complications. Using the SAPNS for biologicalconfinement will reduce post operative complications in endoscopic andopen surgical procedures. Efficacy has been demonstrated on brain,spinal cord, gastrointestinal tract, liver, muscle, arteries and veins.

For example, a partial resection is currently performed as follows. Thesurgeon performs a partial resection of the intestine to remove aprecancerous area. The incision is made and the intestines are gentlylifted out of the intraperitoneal cavity and placed on the table next tothe patient. The offending area is resected and the two ends of theintestine are then ligated together. Before the intestines are put backin the body there is a colostomy bag connected to the upper end of theintestine and the area of the operation is disinfected. The intestinesare replaced in the abdomen and are sewn back up. A drain is placed inthe abdomen to make sure there is no leakage or bleeding. In contrast,using the self-assembling peptide material, a partial resection isperformed as follows. The doctor opens the abdomen and finds theoffending part of the intestine. It is isolated with additional liquidthat is poured into the intraperitoneal cavity to isolate it from therest of intraperineal cavity. The surgeon reaches through the gel thatwas formed by the liquid and resects the intestine. The two ends areligated together and the area is checked for any changes in color.Because the gel also has an indicator die that changes to blue if thereis any leakage of gastric fluids or bacteria. All of the blue is removedwith suction. A little more material is sprayed around the area of therepair and the abdomen is sewn up.

In some cases, the materials described herein may be useful in thetreatment of scars. Experiments have demonstrated that application ofthe self-assembling material can be used to block formation of scarringin the central nervous system (CNS). Administration of the materials atthe site of lesion blocks the stable formation of a scar, which canpermit regeneration through that site. Removing the scar that developsin the central nervous system (CNS) permits axons to grow across theinjury site.

In some cases, the materials may comprise a chelating metal forchelation-enhanced wound healing: The material can be used for thedelivery of a chelator such as iron to a site so it can be used by thebody in the local environment to rebuild basement membrane. For example,in tissues that do not contain enough iron, the delivery of iron in astable form will help healing and the rebuilding of tissue. In somecases, the material comprising a chelating metal may reduce or preventinflammation. In some embodiments, metals associated with a cystine orcystine-like residue can be incorporated in the material so there islittle or no steric hindrance with the assembly of the matrix in-vivo orin-vitro.

The materials described herein can be used to create a clean localenvironment to perform surgery; isolate structures and migration ofcontaminates; inflate structures for surgical procedures, i.e.intestine; surround structures that are being removed that may leak,i.e. appendix, patch holes in body; allow for surgery in dirtyenvironments; used with scope procedures to surround the organ beforethe operation to contain any leakage; used to create a barrier toprevent adhesions while performing abdominal surgery; and used for forma gasket between the scope and the insertion point of the scope.Benefits during surgery are that the material is optically clear, has along shelf life at room temperature, can be operated through it,shortens prep time, eliminates counting sponges, isolates each structurein the surgical field, shortens clean up time of the operating room,shortens surgical time, reduces or eliminates cross contamination causedby other irrigants, the material is biocompatible, the breakdownproducts are natural and are absorbed by the body. The material is easyto manipulate, can be injected at the location needed, should eliminateStaphylococcal infections, may be able to reduce the cost of surgicaltheater disposables paper, reduce biohazard bags since the material canbe boiled to sterilize after the procedure to yield steam. Since thematerial is clear it should enable the surgeon to operate faster becausethe operating field is clear of blood. The elimination of wound packingto control bleeding could reduce the operating time as much as 50% in acomplicated case. Post-op infection, due to secondary infection, may bereduced by the use of the material since it can coat the wound duringand after surgery, thus reducing contamination from foreign bodies.Post-op care may be able to use the material to reduce infection due todrainage by slowing the spread of particulate material in the abdomen orchest cavity.

While the compositions can be removed from a site of application (e.g.,a bleeding vessel) at any time, a physician may wish to allow them toremain in place even after the initial goal of promoting hemostasis hasbeen achieved in order to promote wound healing.

Where the compositions include self-assembling peptides, those peptidescan include amino acid residues that are naturally occurring and thatcan be absorbed by the body. The compositions are not difficult tomanipulate, and they can be easily dispensed on an as-needed basis.Their features (e.g., stiffness) can be altered readily by altering theconcentrations of components therein (e.g., by altering theconcentration of self-assembling peptides in a given composition). Asthe resulting, assembled structure does not significantly impair one'sview of an underlying tissue, and does not have to be removed before aprocedure can be carried out. For example, a physician can assess a burnor other surface trauma that has been treated in the field with acomposition described herein. In the operating room, a surgeon can makean initial incision through the material and can continue to operatewith standard equipment, such as scalpels and clamps, or more modernmeans, such as lasers, in an internal field to which the compositionsmay also have been applied. Another advantage may be realized in time,as use of the compositions can decrease the time required to prepare apatient for surgery. As the compositions can be applied around the siteof an incision and form a coating to protect against infectious agents,there is less need to shave a patient's skin, apply drapes, and applydisinfectants.

Given the structural integrity of the assembled scaffolds, they can beremoved from an area in which they have formed if desired. Thus, anassembled scaffold can be removed by, for example, suction, or bylifting it away with an instrument such as forceps, or wiping it awaywith a swab or gauze. For example, the scaffold can be removed afterhemostasis is achieved or in the course of cleaning a wound. Based onstudies to date, the scaffold or a majority thereof can be removedwithout damaging the underlying tissue. Where the assembled scaffoldsare formed ex vivo, they can be removed from a mold and usedsubsequently (e.g., implanted in a tissue or tissue void). Thecompositions should reduce the amount of material that requires disposalor cleaning afterward (e.g., surgical drapes, sponges, and otherbiohazards).

“Nanodrapes” can be used to replace traditional paper or cloth drapes,by limiting infection following application directly to the patient, forexample, by spraying or otherwise coating the patient or the area aroundthe surgical incision. Currently a patient is prepared for surgery byshaving, scrubbing, disinfecting and draping after positioning on thesurgical table. Then bactericide and tape is applied to the area wherethe surgery is to be performed. The self-assembling composition can beapplied in place of drapes by spraying the warm liquid onto the bodywhere it self-assembles into a thin second skin. This material has apore size that is smaller than any bacteria can fit through, so itprotects from any airborne contaminants, and because the one millimeterthick material can contain a mild anti-bactericide, that clings to thebody like a second skin. The material can also have a hydratingcomponent for the skin so it does not dry out. There is no worry aboutgetting the material into the wound site because it will be broken downby the body. Color can be added to it so it is easier to determine if itis all washed off after the operation.

A scaffold (e.g., a nanoscale structured material) can be provided byintroducing, to a subject, a precursor of the scaffold at a location, orin the vicinity of a location, where the scaffold is desired (e.g., tocontrol movement or leakage of a bodily substance, to protect a wound,or to promote tissue repair). Precursors (e.g., self-assemblingpeptides) are provided in the vicinity of a location when they areprovided at a position that is close enough to the targeted area (e.g.,a bleeding vessel, a diseased section of the digestive tract, or an areaof burned skin) that they reach the targeted area in an effectiveamount. The precursors, which may be homogenous or heterogeneous (e.g.,one may apply a single type of self-assembling peptide or a mixture oftwo or more different such peptides), can be contained within acomposition and, upon contact with physiological conditions, assemble toform the scaffold (e.g., a nanoscale structured material). Thus, theprecursors can assemble in situ (i.e., within the body of a subject ator in the vicinity of administration).

The nanoscale structured material may include, or its assembly mayinvolve, additional components present in situ, (e.g., ions). Thus,precursors such as self-assembling peptides can be applied in a solutionthat is substantially free of ions (e.g., substantially free ofmonovalent cations) and self-assemble to form a macroscopic structurewhen they come in contact with such ions in the body (e.g., in a bodilysubstance such as blood, gastrointestinal contents, and the like). Forexample, a solution containing precursors can be applied at, or in thevicinity of, a site of gastric or intestinal perforation or a site wherea surgical incision has been or will be made.

The scaffold can also be provided in the form of a gel, as theprecursors (e.g., self-assembling peptides) can be assembled prior tointroducing a composition to a targeted area (e.g., the site at which anincision will be made for a surgical procedure). The assembled structuremay assume any convenient shape.

The scaffold can also be provided by providing precursors in the form ofa dry powder. A “dry” powder will have a relatively low liquid content(e.g., sufficiently low that the particles therein are readilydispersible). Self-assembling peptides provided in the form of a drypowder will assemble when they come into contact with a bodily fluidcontaining monovalent cations, and a solution containing such ions maybe added if desired to alter the rate at which the scaffold forms or itsstiffness. Self-assembling peptides may be provided as emulsions or, asdescribed above, molded into preformed shapes that can be inserted intoa body cavity or wound site in a manner similar to the manner thatsurgical sponges are currently-used. If desired, a binder can be addedto a dry powder which is then formed into a desired shape. Regardless ofthe precise manner in which the scaffold is assembled (e.g., whether bybringing a liquid formulation containing precursors into contact withthe body or a dry powder into contact with an ion-containing solution exvivo), the formed scaffolds can assume a desired shape. Where the sizeand shape is such that the scaffold fills the lumen of a blood vessel,the scaffold can be used a vascular plug.

A preventative measure can be carried out before a subject experiencesan unwanted event (e.g., before an injury occurs or before bleedingbegins). Thus, the site of administration can be a site of potentialmovement or potential leakage, and the application can be made toprevent or minimize such movement or leakage should it occur. When usedin the context of a therapeutic procedure or treatment, the compositionscan reverse, alleviate, or inhibit the progress of a condition (e.g., astate, syndrome, disease, or a sign, symptom, or manifestation of such).Methods of treating a subject are generally carried out once the subjectis recognized as having a condition amenable to treatment, and any ofthe methods described herein, whether best described as prophylactic ortherapeutic, can include a step of identifying an amenable subject.

As the compositions described here can be used to inhibit movement of abodily substance in a subject, including movement within or from theepidermis, the compositions can be employed in the context of performingsurgery and may be described as new methods for performing surgery orgenerating a surgical field. The methods, whether performed in thecontext of surgery or not, can include a step of identifying a subjectin need of treatment and a step of providing a nanoscale structuredmaterial, or a precursor thereof, at or in the vicinity of a site whereunwanted movement has occurred or is expected to occur. For example, onecan identify a patient who is about to undergo a surgical procedure andprovide a biocompatible composition comprising self-assembling peptidesand a vasoconstrictor, a coloring agent, or a local anesthetic agent toa site at which an incision or other invasive maneuver will be made orhas been made. The bodily substance that is affected may be a fluid suchas blood or a blood product, serous exudate (an inflammation-associatedexudate composed largely of plasma, which typically appears as a clearor amber-colored fluid), pus, gastric juice, urine, bile, cerebrospinalfluid (CSF), pancreatic juice, and the like. The bodily substance may beviscous, sludge-like or semi-solid but will generally exhibit an abilityto flow or move. Substances of this nature include the contents of thegastrointestinal tract. The composition may be removed after application(e.g., after hemostasis is achieved or an operation on the bowel iscomplete) or may be left in place. For example, the compositions can beapplied to accelerate hemostasis or inhibit movement of intestinalcontents during surgery and some or all of the scaffold may be left inplace when the operation is complete. This provides a substantialadvantage relative to the use of sponges and other materials that mustbe removed prior to closure. The compositions can be removed in avariety of ways (e.g., by wiping or by suction).

The compositions can also be applied to shield an underlying area (e.g.,an area of burned or otherwise injured skin or other tissue) and can,therefore, help to prevent contaminants (e.g., foreign substances) fromcoming into contact with the area (i.e., the compositions can be used asa barrier or shield). A physician or other health-care provider canexamine a wound through the material, and a surgeon can operate throughit, while it is in place. Contaminating substances that have landed onthe material during the procedure could then be removed by virtue ofremoving the material.

The compositions can be administered to stabilize a wound prior todefinitive treatment (e.g., while the victim is awaiting transport to ahospital or during transit). The compositions are similarly useful whereoperations are conducted under conditions of less than optimal sterility(e.g., in field hospitals or in areas of the world where access tosterile operating rooms is limited). The compositions and methods havethe potential to significantly reduce the likelihood of contamination ininstances such as these. The self-assembling peptide material can alsobe locally applied in combination with anesthetic in the local areawhere a procedure is to take place and can be applied at a higherconcentration to reduce organ movement during surgery. This may reducecognitive deficits to older patients by reducing the general anestheticload. A thin layer can be sprayed on the tissue or skin where thesurgeon is operating. It can be applied separately or together,administering specific anesthetic for specific organs. Skin hasdifferent receptors than intestines and the need for a specificanesthetic is needed for each of the organs. Intestines need to stopmoving during surgery while the blood and blood vessel contraction needto remain constant.

Treatment and prevention of bleeding: Any individual who has anincreased risk of suffering undesirable bleeding, which may or may notbe excessive or immediately life-threatening, can be treated with thecompositions described herein. These individuals include those withblood clotting disorders such as hemophilia, patients who are receivinganticoagulant therapy, patients who suffer recurrent nosebleeds, andindividuals undergoing surgery, particularly major surgery or proceduresthat involve accessing an artery. Without limitation, the surgery orprocedure can be an operation on the nervous system, eye, ear, nose,mouth, pharynx, respiratory system, cardiovascular system, digestivesystem, urinary system, musculoskeletal system, integumentary (skin)system, or reproductive system. Specific examples of surgeries andprocedures in which the compositions can be used include arteriography,angiocardiography, cardiac catheterization, repair of obstetriclaceration, removal of coronary artery obstruction, insertion of stent,Caesarean section, hysterectomy, reduction of fracture, coronary arterybypass graft, cholecystectomy, organ transplant, total joint (e.g.,knee, hip, ankle, shoulder) replacement, appendectomy, excision ordestruction of intervertebral disk, partial excision of the largeintestine, mastectomy, or prostatectomy.

Accident victims, individuals engaged in combat, and women giving birthare also at risk of experiencing significant blood loss. Thecompositions can be applied to a site of obstetric bleeding (e.g.,within the uterus, vagina, or neighboring tissue) in order to acceleratehemostasis. For example, the compositions can be applied to a placentaltear or used to pack the uterus to control bleeding. As with otherindications, compositions applied to the reproductive tract can beremoved or left in place. Spontaneous hemorrhage, aneurysm rupture,esophageal varices, gastric ulcers, ulcers of the upper portion of theintestine (e.g., duodenal ulcers) are also medical conditions in whichconsiderable bleeding can occur, and these individuals can also betreated as described here.

The precise source of the bleeding can vary and can be from any bloodvessel in the arterial or venous system (e.g., an artery, arteriole,capillary or capillary bed, venule, or vein). The size of the vessel mayrange from large (e.g., the compositions can inhibit bleeding from theaorta, the iliac or femoral artery, or a portal vein) to small (e.g., acapillary), and the vessel may be located anywhere in the body (e.g., ina solid organ such as liver, the stomach, intestine, skin, muscle, bone,the lungs, or the reproductive system).

The time normally required for blood clotting can be prolonged whenplasma levels of clotting factors and/or platelets are low or in casesin which an individual has received an anticoagulant (e.g., warfarin orheparin). Bleeding frequently persists for considerably longer than theaverage clotting time when there is more than minimal damage to bloodvessel integrity. Based on the studies, it is expected that thecompositions will cause hemostasis in a period of time that is lessthan, and in at least some cases much less than, the average bloodclotting time. Although the compositions are not limited to those thatachieve hemostasis in any given time (and uses such as protecting anarea from contamination or promoting tissue healing are independent ofthis function), the compositions may confer a benefit to a bleedingsubject in as little as five seconds following application. Othercompositions can exert an effect in about 10, 15, or 20 secondsfollowing application. The effective period can be characterized in amanner other than absolute time. For example, compositions may reducethe time required to achieve hemostasis by between 25% and 50%; between50% and 75%; or between 75% and 100% relative to the time required wheniced saline is applied. The time required to achieve hemostasis can bereduced by approximately 2-, 3-, 4-, or 5-fold relative to the timerequired when iced saline is applied.

The peptide concentration may be selected with reference to variablessuch as the caliber of the vessel, the extent to which it has beeninjured, and the force with which blood is exiting (or would exit uponinjury). Higher peptide concentrations will be desirable to promotehemostasis from a major vessel (e.g., the aorta, brachiocephalic,carotid, subclavian, celiac, superior mesenteric, renal, iliac, femoral,or popliteal arteries). Useful concentrations can range from betweenapproximately 0.1-10% (e.g., 1-10%; 0.5-5%; 14%; 0.1-2%; 0.1-3%; 0.1-4%;0.1-5%; and 1-8% (e.g., about 1, 1.5, 2, 2.5, 3, 4, 5, 6, or 7%). Anysubrange, or any specific value within any of the aforesaid ranges, canbe used. Any of the aforementioned concentrations may also be used forthe other indications described herein.

As noted, bleeding can be due to any of a large number of differentcauses and can be internal or external. The compositions can be appliedregardless of the cause or the nature of the cause (e.g. whether causedby a disease process or intentional or accidental trauma). Thecompositions can be used to achieve hemostasis in a confined space(e.g., inside a hollow organ) or at or near the body's surface. Forexample, the compositions can be applied to a partly or completelysevered body part such as a limb or digit. In that event, thecompositions may be serving multiple functions; they may not onlypromote hemostasis, but also protect the wounded tissue fromcontaminants and promote tissue healing. More specifically, thecompositions can be applied to a wound, left in place for a period oftime sufficient to achieve hemostasis and for blood clotting to occur,and then removed. Contaminating material such as particulates andinfectious agents adhered to the peptide gel would be removed with it. Asterile dressing may then be applied. Of course the compositions can beapplied for purposes of cleaning a wound, preventing contamination, orpromoting tissue healing even after hemostasis has been achieved or insituations in which acceleration of hemostasis is not needed.

When used to treat a nosebleed, the compositions arc inserted into theappropriate nostril and can be left in place until the bleeding hassubsided. The compositions can be easily removed by suction (e.g., usingan eyedropper or syringe) or may be removed by other physical means,including simply blowing the nose.

The compositions can also be left in place on a wound, and a dressingcan be applied over the composition. Since the composition itself iseasily removed, its presence under the dressing can help prevent thedressing from sticking to the damaged tissue. If desired, a bandagehaving a transparent portion may be used so the injured site can beviewed through the transparent portion of the bandage and the peptidestructure below. This would allow a physician to monitor the progress ofthe healing without removing the dressing. Modified bandages aredescribed further below and are within the scope of the presentinvention.

Many medical procedures involve vascular puncture, which can be followedby significant bleeding. A self-assembling peptide composition can beapplied to the wall of a punctured vessel, e.g., during withdrawal of aninstrument used to puncture the vessel. A vascular plug formed fromself-assembling peptides provides an alternative to existing vascularplugs and devices such as those described in U.S. Pat. Nos. 5,192,302;5,222,974; 5,645,565; and 6,663,655. The vascular plug can be formed insitu (e.g., at a site of vascular puncture), or can be preformed andapplied to the site.

More generally, compositions comprising nanostructured materials orprecursors thereof (e.g., self-assembling peptides) can be used forsealing any passage through tissue. The present methods thereforeinclude methods of sealing a passage through tissue by applying acomposition comprising a nanoscale structured material (e.g.,self-assembling amphiphilic peptides) to one or both ends of the passageor to its interior. The tissue can be, for example, the wall of a bloodvessel, the wall of an organ, subcutaneous tissue, or adipose tissue.Sealing the passage can result in hemostasis. The passage can also be afistula (i.e., an abnormal connection between two organs or bodystructures or between an organ or structure and the external world). Ifdesired, a surgeon can apply the compositions to the interior of atubular structure such as the intestine or a blood vessel, resect andligate the intestine or blood vessel in the gel, and evacuate the gelfrom the interior of the structure to restore continuity of thestructure and allow reperfusion of the area with blood or other bodysubstances.

For surgical applications, the wound or any part of the surgical fieldcan be packed with a composition comprising self-assembling peptides.This approach can be used instead of wound packing as it isconventionally performed during surgery. As the compositions containbiocompatible and biodegradable material, they can be left in place,thereby avoiding the need for removal at the end of the procedure andavoiding the need for a subsequent operation for this purpose.Biodegradable materials can be broken down physically and/or chemicallywithin cells or within the body of a subject (e.g., by hydrolysis underphysiological conditions or by natural biological processes such as theaction of enzymes present within cells or within the body) to formsmaller chemical species which can be metabolized and, optionally,reused, and/or excreted or otherwise disposed of. Preferably, thebiodegradable compounds are biocompatible.

Gastrointestinal bleeding, which can occur as a consequence of ulcers orangiodysplasia, is a relatively common and serious condition that can befatal if left untreated. Bleeding esophageal varices, and bleedinggastric or duodenal ulcers can be particularly severe. A number ofendoscopic therapeutic approaches have been developed to achievehemostasis, such as the injection of sclerosing agents, the attachmentof mechanical hemostatic devices, and contact electrocautery techniques.The compositions can be administered at, or in the vicinity of, an ulceror a site of bleeding in the esophagus, stomach, small intestine, orlarge intestine. Bleeding in the distal portion of the large intestine,rectum, or anus (e.g., hemorrhoids) can also be treated in this manner.

Rupture of an aneurysm can represent a catastrophic event with rapidlyfatal consequences. Ruptured aortic aneurysms can rapidly result inexsanguination despite prompt medical attention. Ruptured intracranialaneurysms frequently have devastating consequences. The compositions andmethods of the invention can be used to treat bleeding from a rupturedaneurysm in an essentially similar manner to the way in which they areused to treat bleeding due to other causes (e.g., by application ofself-assembling precursors or a preformed structure to the site ofbleeding). Given the often severe consequences of aneurysm rupture,surgical repair is often attempted. The compositions can be applied inthe context of any attempted repair (e.g., during open surgery orendovascular repair (e.g., with placement of a graft and/or stent)).More specifically, the present methods include treating an aneurysm byintroducing a composition comprising a nanoscale structured material orprecursor thereof (e.g., a composition comprising self-assemblingpeptides) into the aneurysm (e.g., into the aneurysm sac). Once anybleeding is under better control, the aneurysm may then be repairedusing any suitable technique. Presence of the peptide structure withinthe aneurysm sac reduces the chance of leakage or rupture prior to orduring these other procedures. The scaffold can be left in place.

Inhibiting movement or leakage of cerebrospinal fluid (CSF): The duramater is the tough, outermost, fibrous membrane that covers the brainand spinal cord, and lines the inner surface of the skull. Leakage ofCSF is a significant complication following injury, surgery, or otherprocedures in which the dura mater is penetrated, including inadvertentpenetration in the course of administering an anesthetic to the epiduralspace. Such leakage can lead to serious sequelae, such as severeheadaches, infection, and meningitis. The composition can inhibitmovement or leakage of CSF in a subject in need thereof afterapplication at, or in the vicinity of, a site of unwanted movement orleakage of CSF. The compositions can be applied over sutures followingdura mater surgery to help prevent CSF from leaking out of the incisionsite. The compositions can also be used to inhibit movement or leakageof fluid from the ear drum.

Inhibiting leakage of contents of the gastrointestinal tract: Thecompositions can inhibit the movement of gastrointestinal contents. Forexample, the structures can prevent leakage of gastrointestinal contentsfollowing gastric or intestinal perforation or during surgery (seeExample 4). The structures can be used to isolate such bodily substancesand prevent their spread within the peritoneal cavity, therebyminimizing contamination and the risk of subsequent chemical peritonitisand/or infection. Gastric contents, which contain digestive secretionsof the stomach glands consisting chiefly of hydrochloric acid, mucin,and enzymes such as pepsin and lipase, can cause injury and/or infectionif released into the peritoneal cavity. Release of intestinal contentsinto the peritoneal cavity represents a frequent event during surgery onthe intestine and can also occur in cases of intestinal perforation or aruptured appendix. The composition can be used to inhibit leakage ofgastrointestinal contents into the peritoneal cavity. The site ofmovement can be a site of gastric or intestinal damage caused by adisease process or a surgical incision. The compositions can be appliedto the exterior of any organ in the digestive system (e.g., the stomach,or small or large intestine) or can be injected or otherwise introducedinto their interior. The compositions can be administered in the courseof resecting a segment of the intestine. For example, one can fill asegment of intestine that extends from a first point to a second pointwith a present composition and resect a portion of the intestine thatlies between the first and second points.

In a related method, one can use the compositions to remove intestinalcontents that have been released into the peritoneal cavity. The methodincludes applying a liquid composition to the released intestinalcontents, allowing the liquid composition to undergo a phase transition,and then removing the gel-like or semi-solid composition. These stepscan be repeated once or more until the surgeon is satisfied with theamount of intestinal contents that have been removed from the peritonealcavity.

One can similarly inhibit movement of the contents of other internalorgans (e.g., organs in the biliary or urinary systems). For example,one can inhibit movement of bile, pancreatic juice (i.e., secretions ofthe exocrineportion of the pancreas that contain digestive enzymes), orurine and/or decontaminate or clean an area into which bile, pancreaticjuice, or urine have been released by application and subsequent removalof the compositions to the site. The methods thus have broad applicationto surgeries for repairing or otherwise treating intestinal, biliary,and/or urinary system defects.

Wound healing: Studies also indicate that the compositions have theability to enhance healing, particularly of an epithelial layer ormuscle, and can therefore be administered to treat a site of tissuedamage. For example, one can apply a composition includingself-assembling peptides to the site of tissue damage. The compositionsappear to both increase the rate of tissue repair and inhibit formationof scar tissue. The compositions can be used for either acute or chronicwound care. For example, they can be applied to skin wounded in anymanner (e.g., lacerated or burned) and to lesions such as diabeticulcers and pressure sores.

These materials can be used to maintain hydration and nutrition topatients that have had burns or in cases the outer skin has beenbreached due to abrasion or burn. In another case the material can beused to maintain body temperature when the patient is covered with thematerial by means of external heat or cooling source.

The materials may be incorporated into various delivery methods,devices, and kits. A variety of devices can be used to introduce thecompositions to a target area of the body. The devices can be simple,such as a syringe, and such devices can be provided together with thecompositions in kits. The composition can be locally delivered at ornear a target area in the body by injection (e.g., using a needle andsyringe), or with a catheter, cannula, or by dispensing (e.g., pouring)from any suitably-sized vessel. The compositions can be delivered withthe assistance of imaging guidance (e.g., stereotactic guidance) ifnecessary. Alternately, a material can be wetted with the compositionand then used to apply a composition to an area of tissue.

For storage and shipping, self-assembling materials can be dissolved ina suitable solvent (e.g., an aqueous medium such as sterile water, andstored for long periods of time prior to use). Peptide-containingsolutions have been stored for up to two years without substantial lossof activity. If partial self-assembly occurs after a prolonged period oftime, physical agitation (e.g., sonication) can be used to restore thematerial to a more liquid state prior to administration. Alternatively,the material can be applied as a gel. If desired, a small amount of ions(e.g., monovalent cations) can be added to a solution prior toapplication. This may speed the process of gel formation. Alternately,monovalent cations can be applied after the solution has beenadministered.

Kits containing syringes of various capacities or vessels withdeformable sides (e.g., plastic vessels or plastic-sided vessels) thatcan be squeezed to force a liquid composition out of an orifice areprovided. In one embodiment, the syringe or vessel contains multiplecompartments, one containing monovalent ions, and the otherself-assembling peptides, which are mixed at the time of administration,through a common needle. An endoscope can be used to deliver thecompositions for treatment of a hollow organ (e.g., the esophagus,stomach, intestine, etc.) or body cavity (e.g., during minimallyinvasive surgery). Minimally invasive surgery refers to an approach tosurgery whereby operations are performed with specialized instrumentsdesigned to be inserted through small incisions or natural bodyopenings, often performed with endoscopic visualization. Examplesinclude laparoscopic surgery, arthroscopic surgery, and endovascularsurgery. An endoscope is typically a long, flexible tube-like device. Inaddition to allowing visualization of internal structures, manyendoscopes have additional diagnostic (e.g. biopsy) and therapeuticcapabilities (e.g. delivery of therapeutic agents) through specialchannels. Colonoscopes, sigmoidoscopes, bronchoscopes, cystoscopes, andlaparoscopes, are variants of an endoscope having features making themparticularly well suited for viewing certain organs, structures, orcavities. Any of these devices can be used to deliver the compositions.Kits may be packaged including an endoscope and a vessel containing asolution comprising self-assembling peptides. Suitable endoscopes areknown in the art and are widely available. Endoscopes are currently inuse to deliver sclerosing agents to sites of esophageal bleeding.

Kits can include self-assembling peptides and one or more of: a syringe,a needle, thread, gauze, a bandage, a disinfectant, an antibiotic, alocal anesthetic, an analgesic agent, surgical thread, scissors, ascalpel, a sterile fluid, and a sterile vessel. The peptides can be insolution or dry (e.g., as a dry powder). Components of the kit may bepackaged individually and are sterile. The kits are generally providedin a container, e.g., a plastic, cardboard, or metal container suitablefor commercial sale. The kit may be styled as a “first aid kit,” inwhich case it will typically have a symbol such as a red cross on theexterior. Any of the kits can include instructions for use.

EXAMPLES Example 1

The following example describes a self-assembling peptide material andits use in the acceleration of hemostasis in the brain.

Complete transection of a branch of the superior sagittal sinus in thebrains of rats and hamsters was performed after removing a portion ofthe skull overlying the transected tissue. Animals were anesthetizedwith an i.p. injection of ketamine (80 mg/kg) and xylazine (8 mg/kg).All surgical procedures were conducted under an operating microscope.Twenty-two animals, including 10 adult hamsters and 12 young adultfemale Spraque-Dawley rats (200-250 g), were treated with either icedsaline or 20 μl of a 1% peptide solution at the site of the sinus branchtransection. The material was prepared by dissolving RADA 16-1(n-RADARADARADARADA-c; SEQ ID NO: 1) peptide in sterile water, and thepeptide-containing solution was applied to the injured tissue with a 31gauge needle attached to a 2 cc syringe.

The experiment was videotaped with a time stamp and was replayed oneframe at a time to evaluate the length of time required for the peptidesolution to form a gel, which effectively impeded bleeding. Hemostasiswas assessed visually, and “complete hemostasis” was defined as thecomplete lack of movement of blood from the wound site. Completehemostasis was achieved within 10 seconds of the application of thepeptide solution in all cases.

A series of pictures was taken of an adult rat in which a portion of theoverlying skull was removed and one of the veins of the superiorsagittal sinus was transected and then treated with a peptide-containingsolution. The initial picture shows the exposed brain and veins of thesuperior sagittal sinus; the next picture shows the cutting of the vein;the next picture shows bleeding from the ruptured vein; and the finalpicture shows the same area five seconds after the peptide solution wasapplied. Complete hemostasis was achieved.

Durations were measured from the start of application of peptidesolution to the completion of hemostasis after transection of the veinsleading to the sinus in the brains of adult rats. Complete hemostasiswas achieved in an average of 8.3 seconds. In the saline controls,cessation of bleeding was never achieved. The saline control experimentwas terminated at the same time point in order to prevent the animalsfrom bleeding to death.

Similar results have been obtained following complete transection of thesuperior sagittal sinus. A higher concentration of peptide (e.g.,approximately 3%-4%) was used in the latter experiment in order toachieve hemostasis. The three saline control cases continued to bleedafter 20 seconds. In the control animals, the iced saline was removedand the peptide solution was applied, resulting in complete hemostasisalmost immediately.

A total of 22 rats and 64 hamsters have been subjected to experiments inwhich peptide-containing solutions effectively achieved hemostasiswithin 10 seconds following application to a site intracranial bleeding.

Example 2

The following example describes a self-assembling peptide material andits use in the acceleration of hemostasis following femoral arterytransection.

The sciatic nerve and the adjacent femoral artery were exposed in adultrats, and the femoral artery was transected. Twelve rats were treated byapplication of 20 μl of a 1% solution of RADA 16-I peptide to the siteof transection using a glass pipette attached to a syringe body, whilecontrols were treated by applying cold saline to the site oftransection. In all treated cases, hemostasis was achieved in less than10 seconds. The saline control cases continued to bleed until theexperiment was terminated at 110 seconds. In these control animals,subsequent replacement of the cold saline with the peptide solutionresulted in almost immediate achievement of complete hemostasis. Aseries of pictures was taken in an adult rat in which the femoral arterywas transected. In the picture taken first, the sciatic nerve and thefemoral artery are exposed. The next picture shows the cutting of theartery, and the next picture shows bleeding. After about five seconds,complete hemostasis was observed in the area of a clear gel formed bythe assembled peptides in the presence of blood and plasma. Theassembled material can be suctioned off the site easily if desired.Complete hemostasis was maintained for the duration of the test (1hour).

Complete hemostasis was achieved in less than 10 seconds. In the salinecontrols, hemostasis was never reached.

Muscle trauma experiments showed immediate hemostasis after 1-2 cmincisions were made in the muscle on the back of a rat. Thespinotrapezius muscles on the back of the rats were exposed and a deepcut was made in the muscle, after which 1% peptide solution (RADA16-I)was applied in the cut. Within 10 seconds, all bleeding had stopped.With the application of iced saline alone, control animals continued tobleed after 20 seconds.

This procedure was duplicated in the muscle of the hind limb(porteocaudalis and musculus tibialis cranialis) and similar resultswere obtained. Between 1% to 100% peptide (RADA 16-1) was applied tolimb wounds, and hemostasis was achieved in all cases. However when anartery or vein was transected 2% or higher material was needed to bringabout hemostasis. With the application of iced saline alone, controlanimals continued to bleed after 20 seconds.

Example 3

The following example describes a self-assembling peptide material andits use in the acceleration of hemostasis in a liver.

To further demonstrate the ability of peptide-containing structures tohalt bleeding of a vessel having relatively low pressure, theintraperitoneal cavity of an adult rat was opened, the liver wasexposed, and the lobus sinister lateralis received a rostral to caudalcut completely transecting a portion of the liver. Profuse bleedingensued. A 1% peptide solution (RADA16-1) was applied to the cut and inits vicinity using a 27 gauge needle and 4 cc syringe. All bleedingstopped within 10 seconds. A series of pictures was obtained. The firstshows exposure of the liver; in the second, the liver is separated, andprofuse bleeding is evident; and in the third, the two portions of theliver are allowed to come back together, and the bleeding continues.After treating the site with 1% peptide solution (applied topically andin the cut), all bleeding stopped within 10 seconds. A clear area wasobserved between the two halves of the lobus sinister lateralis. Thisprocedure was repeated several times with the same result.

A similar experiment demonstrated the ability of the peptide structuresto halt bleeding of a vessel in the liver having a higher pressure. Aseries of pictures illustrate the experiment. The first depicts theopened intraperitoneal cavity and exposed liver; in the second, thelobus sinister lateralis received a transverse cut completelytransecting a portion of the liver and a major branch of the portalvein; and the third shows profuse bleeding from the site of injury. Thecut was treated with 4% peptide solution applied topically and in thecut. All bleeding stopped within 10 seconds. The lower part of the lobussinister lateralis was pulled downward to show that the peptidestructure is in the cut. The site did not bleed even when subjected tothis physical stress. Ten minutes later, there was still no bleeding.Thus, application of 4% peptide solution brings about completehemostasis in a high pressure bleeding environment in less than 10seconds. Treatment with a 2% or 3% peptide solution was tested in thesame type of experiment and complete hemostasis was also achieved.Treatment with a 1% solution resulted in partial cessation of bleeding.In addition, 30 seconds after treatment the excess peptide structure waswiped away from the injury site and hemostasis was maintained. Thisprocedure was repeated several times with the same result.

In other experiments ¼ of the lobe in the lower right quadrant of thelobus sinistras laterialis was removed, and the margin was treated witha topical application of 2% peptide (RADA16-I) to the site of injury.Bleeding stopped in less than 10 seconds. One minute later the peptidewas removed, and complete hemostasis was achieved at the margin of theliver.

Example 4

The following example describes a self-assembling peptide material andits use in reducing the flow of gastric fluid in an intestine.

The intestine of an adult rat was perforated with a small cut at thelevel of the duodenum that resulted in the leakage of gastric fluid intothe intraperitoneal cavity. When the site was treated with 2% peptide(RADA16-I) solution all leakage of gastric fluids from the intestinestopped. An additional volume of 2% peptide solution was injected intothe duodenum at the level of the injury. This prevented all leakage fromthe intestine for one hour, the duration of the procedure. In thecontrol cut at the level of the duodenum, the wall of the intestineinverted and gastric fluids continued to leak from the site of injurywhen left untreated. When the site was treated with peptide solution 15minutes after the injury, the peptide treatment also stopped all leakagefrom this injury site. In addition, the treatment stopped theprogression of the intestinal wall inversion.

Example 5

The following example describes a self-assembling peptide material andits use in accelerating the healing of skin wounds.

To demonstrate the ability of the self-assembling peptides to enhancewound healing, animals were subjected to punch biopsies of the skin andsubcutaneous tissue. The regions from which the biopsies were taken wereeither treated by a single application of self-assembling peptide(RADA16-I) solution or were left untreated. The wounds were leftunbandaged. A series of pictures of a 4 mm punch biopsy healing test inwhich injured animals were treated with the self assembling peptide andcompared to matching cases with no treatment illustrates the results.The wounds were photographed on day 0, day 1, day 4, and day 7. Thetreated wounds healed much faster as evidenced by the contraction of thewound site in all three punches as early as day 1. Treatment with thepeptide appeared to speed healing by as much as 5 days in some cases. Inall cases, shrinkage of the wound site happened faster in the treatedcases.

Example 6

The following example describes the use of compositions containinglidocaine.

RADA 16 mixed with lidocaine and the mixture was applied to the skin ofadult rats before applying a pin prick. It is a 5% mix of lidocaine andRADA1-16. Applied on the skin and left for the duration of the testing.When mixed with a self-assembling peptide, the response to pin prick wasmuted four times longer than the response was muted using lidocainealone. In addition, we applied solutions of self-assembling peptides andlidocaine to the intestines of two rats while performing intestinalsurgery. The solution reduced peristalsis for the duration of thesurgery with no apparent side effects to the animals.

The foregoing description is to be understood as being representativeonly and is not intended to be limiting. Alternative systems andtechniques for making and using the compositions and devices of theinvention and for practicing the inventive methods will be apparent toone of skill in the art and are intended to be included within theaccompanying claims.

While several embodiments of the present invention have been describedand illustrated herein, those of ordinary skill in the art will readilyenvision a variety of other means and/or structures for performing thefunctions and/or obtaining the results and/or one or more of theadvantages described herein, and each of such variations and/ormodifications is deemed to be within the scope of the present invention.More generally, those skilled in the art will readily appreciate thatall parameters, dimensions, materials, and configurations describedherein are meant to be exemplary and that the actual parameters,dimensions, materials, and/or configurations will depend upon thespecific application or applications for which the teachings of thepresent invention is/are used. Those skilled in the art will recognize,or be able to ascertain using no more than routine experimentation, manyequivalents to the specific embodiments of the invention describedherein. It is, therefore, to be understood that the foregoingembodiments are presented by way of example only and that, within thescope of the appended claims and equivalents thereto, the invention maybe practiced otherwise than as specifically described and claimed. Thepresent invention is directed to each individual feature, system,article, material, kit, and/or method described herein. In addition, anycombination of two or more such features, systems, articles, materials,kits, and/or methods, if such features, systems, articles, materials,kits, and/or methods are not mutually inconsistent, is included withinthe scope of the present invention.

The indefinite articles “a” and “an,” as used herein in thespecification and in the claims, unless clearly indicated to thecontrary, should be understood to mean “at least one.”

The phrase “and/or,” as used herein in the specification and in theclaims, should be understood to mean “either or both” of the elements soconjoined, i.e., elements that are conjunctively present in some casesand disjunctively present in other cases. Other elements may optionallybe present other than the elements specifically identified by the“and/or” clause, whether related or unrelated to those elementsspecifically identified unless clearly indicated to the contrary. Thus,as a non-limiting example, a reference to “A and/or B,” when used inconjunction with open-ended language such as “comprising” can refer, inone embodiment, to A without B (optionally including elements other thanB); in another embodiment, to B without A (optionally including elementsother than A); in yet another embodiment, to both A and B (optionallyincluding other elements); etc.

As used herein in the specification and in the claims, “or” should beunderstood to have the same meaning as “and/or” as defined above. Forexample, when separating items in a list, “or” or “and/or” shall beinterpreted as being inclusive, i.e., the inclusion of at least one, butalso including more than one, of a number or list of elements, and,optionally, additional unlisted items. Only terms clearly indicated tothe contrary, such as “only one of” or “exactly one of,” or, when usedin the claims, “consisting of,” will refer to the inclusion of exactlyone element of a number or list of elements. In general, the term “or”as used herein shall only be interpreted as indicating exclusivealternatives (i.e. “one or the other but not both”) when preceded byterms of exclusivity, such as “either,” “one of,” “only one of,” or“exactly one of.” “Consisting essentially of,” when used in the claims,shall have its ordinary meaning as used in the field of patent law.

As used herein in the specification and in the claims, the phrase “atleast one,” in reference to a list of one or more elements, should beunderstood to mean at least one element selected from any one or more ofthe elements in the list of elements, but not necessarily including atleast one of each and every element specifically listed within the listof elements and not excluding any combinations of elements in the listof elements. This definition also allows that elements may optionally bepresent other than the elements specifically identified within the listof elements to which the phrase “at least one” refers, whether relatedor unrelated to those elements specifically identified. Thus, as anon-limiting example, “at least one of A and B” (or, equivalently, “atleast one of A or B,” or, equivalently “at least one of A and/or B”) canrefer, in one embodiment, to at least one, optionally including morethan one, A, with no B present (and optionally including elements otherthan B); in another embodiment, to at least one, optionally includingmore than one, B, with no A present (and optionally including elementsother than A); in yet another embodiment, to at least one, optionallyincluding more than one, A, and at least one, optionally including morethan one, B (and optionally including other elements); etc.

In the claims, as well as in the specification above, all transitionalphrases such as “comprising,” “including,” “carrying,” “having,”“containing,” “involving,” “holding,” and the like are to be understoodto be open-ended, i.e., to mean including but not limited to. Only thetransitional phrases “consisting of” and “consisting essentially of”shall be closed or semi-closed transitional phrases, respectively, asset forth in the United States Patent Office Manual of Patent ExaminingProcedures, Section 2111.03.

What is claimed:
 1. An in vivo barrier structure, comprising: acomposition comprising self-assembled peptidomimetics comprisingalternating hydrophobic and hydrophilic residues and alternatingpositively-charged and negatively-charged residues, wherein thepeptidomimetics are selected from the group consisting of α-peptides,β-peptides, γ-peptides, and δ-peptides, wherein the α-peptidepeptidomimetics are selected from the group consisting of N,N′-linkedoligoureas, oligopyrrolinones, oxazolidin-2-ones, azatides andazapeptides, and wherein the barrier structure is formed followingcontact of the peptidomimetics with a bodily fluid and/or contaminant ofa tissue.
 2. The structure as in claim 1, wherein the bodily fluid isblood, plasma, interstitial fluid, cerebral spinal fluid, tissue fluids,fluid produced by a parenchymal tissue, serous exudate, pus, gastricjuice, urine, or bile.
 3. The structure as in claim 1, wherein thebodily fluid is blood.
 4. The structure as in claim 1, wherein thecontaminant comprises bacteria, cells, infection, pus, serous exudate,bile, pancreatic fluid, or a substance contained within the stomach,intestine, or urinary tract, or an airborne contaminant.
 5. Thestructure as in claim 1, wherein the structure further comprises atherapeutic agent, a prophylactic agent, a diagnostic agent, apharmaceutically acceptable diluent, filler, or a stabilizing agent. 6.The structure as in claim 5, wherein the structure further comprisesanti-inflammatories, vasoconstrictors, anti-infectives, anesthetics,growth factors, cells, organic compounds, coloring agents, vitamins, ormetals.
 7. The structure as in claim 1, wherein the barrier structurechanges in phase relative to the composition prior to contact with thebodily fluid and/or contaminant.
 8. The structure as in claim 1, whereinthe peptidomimetics self-assemble into a plurality of nanofibers.
 9. Thestructure as in claim 8, wherein each nanofiber of the plurality ofnanofibers has a diameter of approximately 10-20 nm.
 10. The structureas in claim 1, wherein the bodily fluid and/or contaminant of the animalcomprises at least one ionic species.
 11. The structure as in claim 10,wherein the ionic species is a monovalent cation, a divalent cation, ora trivalent cation.
 12. The structure as in claim 11, wherein the ionicspecies is Li⁺, Na⁺, K⁺, or Cs⁺.
 13. The structure as in claim 1,wherein the hydrophilic residues alternate with one positively chargedand one negatively charged residue.
 14. The structure as in claim 1,wherein the hydrophilic residues alternate with two positively chargedand two negatively charged residues.
 15. The structure as in claim 1,wherein the hydrophilic residues alternate with three positively chargedand three negatively charged residues.
 16. The structure as in claim 1,wherein the hydrophilic residues alternate with four positively chargedand four negatively charged residues.
 17. A composition, comprising:self-assembling peptidomimetics, wherein the peptidomimetics comprisealternating hydrophobic and hydrophilic residues, and alternatingpositively-charged and negatively-charged residues, wherein thepeptidomimetics are selected from the group consisting of α-peptides,β-peptides, γ-peptides, and δ-peptides, wherein the α-peptidepeptidomimetics are selected from the group consisting of N,N′-linkedoligoureas, oligopyrrolinones, oxazolidin-2-ones, azatides andazapeptides, and wherein the composition forms a structure followingcontact of the peptidomimetics with a bodily fluid of a tissue andwherein the structure inhibits movement of a bodily fluid and/orcontaminant of a tissue.
 18. The composition as in claim 17, wherein thepeptidomimetics are in solid form.
 19. The composition as in claim 17,wherein the peptidomimetics are contained in solution.
 20. Thecomposition as in claim 18, wherein the solid is a spray; a powder or alyophilized powder; or a plurality of pellets, granules, or particles.21. The composition as in claim 17, wherein the composition furthercomprises a nasal plug, a bandage, a suture, a tape, an adhesive, asurgical drape, a foam, or a matrix.
 22. The composition as in claim 17,wherein the composition is a gel or a foam.
 23. A kit comprising thecomposition as in claim 17, and instructions for use in inhibitingmovement of a bodily fluid and/or contaminant in or out of a wound orsite of injury or surgery of a tissue.
 24. The kit as in claim 23,wherein the kit further comprises a syringe or vessel comprising one ormore compartments, wherein a first compartment comprises the compositionand a second compartment comprises a solution which, when combined withthe composition, dissolves or hydrates the composition to form amaterial that inhibits movement of a bodily fluid and/or contaminant.25. The composition as in claim 17, wherein the structure is formed uponinteraction of the peptidomimetics with at least one ionic speciespresent in or on the tissue.
 26. The composition as in claim 19, whereinthe solution is water.